A nucleic acid strand is inherently
directional, and the "5 prime end"
has a free hydroxyl (or phosphate) on a 5'
carbon and the "3 prime end" has a
free hydroxyl (or phosphate) on a 3' carbon
(carbon atoms in the sugar ring are numbered
from 1' to 5'. That's simple enough for an
RNA strand or for single-stranded (ss) DNA.
However, for double-stranded (ds) DNA it's
not so obvious - each strand has a 5' end
and a 3' end, and the 5' end of one strand
is paired with the 3' end of the other
strand (it is "antiparallel"). One
would talk about the 5' end of ds DNA only
if there was some reason to emphasize one
strand over the other - for example if one
strand is the sense strand of a gene. In
that case, the orientation of the sense
strand establishes the direction.
3' flanking region: A
region of DNA which is not copied into the
mature mRNA, but which is present adjacent
to 3' end of the gene. It was originally
thought that the 3' flanking DNA was not
transcribed at all, but it was discovered to
be transcribed into RNA, but quickly removed
during processing of the primary transcript
to form the mature mRNA. The 3' flanking
region often contains sequences which affect
the formation of the 3' end of the message.
It may also contain enhancers or other sites
to which proteins may bind.
3' untranslated region:
A region of the DNA which is transcribed
into mRNA and becomes the 3' end or the
message, but which does not contain protein
coding sequence. Everything between the stop
codon and the polyA tail is considered to be
3' untranslated (see Figure 4). The 3'
untranslated region may affect the
translation efficiency of the mRNA or the
stability of the mRNA. It also has sequences
which are required for the addition of the
poly(A) tail to the message (including one
known as the "hexanucleotide",
AAUAAA).
5' flanking region: A
region of DNA which is not transcribed into
RNA, but rather is adjacent to 5' end of the
gene. The 5'-flanking region contains the
promoter, and may also contain enhancers or
other protein binding sites.
5' untranslated region:
A region of a gene which IS transcribed into
mRNA, becoming the 5' end of the message,
but which does not contain protein coding
sequence. The 5'-untranslated region is the
portion of the DNA starting from the cap
site and extending to the base just before
the ATG translation initiation codon. While
not itself translated, this region may have
sequences which alter the translation
efficiency of the mRNA, or which affect the
stability of the mRNA.
7-methyl-guanosine
triphosphate: The methylated form of
guanosine triphosphate used to cap an mRNA
molecule in most eukaryotes. (See cap)
A
Ablation experiment:
An experiment designed to produce an animal
deficient in one or a few cell types, in
order to study cell lineage or cell
function. The idea is to make a transgenic
mouse with a toxin gene (often diphtheria
toxin) under control of a specialized
promoter which activates only in the target
cell type. When embryo development
progresses to the point where it starts to
form the target tissue, the toxin gene is
activated, and that specific tissue dies.
Other tissues are unaffected.
Acrylamide gels: A
polymer gel used for electrophoresis of DNA
or protein to measure their sizes (in
daltons for proteins, or in base pairs for
DNA). See "Gel electrophoresis".
Acrylamide gels are especially useful for
high-resolution separations of DNA in the
range of tens to hundreds of nucleotides in
length.
Adapter: A short
chemically synthesized DNA double strand
which can be used to link the ends of two
DNA molecules.
Adenine (A): One of
the purine bases found in DNA and RNA
(6-aminopurine), one member of the base pair
A- T (adenine- thymine).
Agarose gel
electrophoresis: A method to analyze the
size of DNA (or RNA) fragments. In the
presence of an electric field, larger
fragments of DNA move through a gel slower
than smaller ones, producing different
migrating "bands". Usually, these
are visualized by soaking the gel in a dye
(ethidium bromide) which makes the DNA
fluoresce under UV light. This is the gel of
choice for DNA or RNA in the range of
thousands of bases in length, or even up to
1 megabase if you are using pulsed field gel
electrophoresis. (See also electrophoresis).
Alkaline Phosphatase:
An enzyme which catalyzes the hydrolysis of
phosphomonoesters of the 5' nucleotides.
Used to dephosphorylate (remove phosphate
groups from) the 5' ends of DNA or RNA
molecules, to facilitate 5' end-labeling
with 32P added back by T4 polynucleotide
kinase; or to dephosphorylate the 5' ends of
DNA molecules to prevent unwanted ligation
reactions during cloning.
Allele: An allele is
one of the alternative (two or more) forms
of a particular gene inherited separately
from each parent; usually found at the same
locus on homologous chromosomes. Equivalent
genes in the two sets might be different,
for example because of single nucleotide
polymorphisms.
Allele-specific ligation:
Technique permitting discrimination of two
allele at locus by providing two short
synthetic oligonucleotides that would bind
ajjacent to teach other on amplified DNA
fragment, and would be ligated in presence
of DNA ligase; if one of alleles containing
mutation overlapped by 3'-end of one
oligonucleotide, its ligation to
oligonucleotide bound 3' to it would be
prevented; to deduce identity of unknown
allele, differentially labelled
oligonucleotide pairs may be designed for
each allele and their ligation efficiency
compared in presence of unknown allele.
Allele-specific PCR
(AS-PCR): Refers to amplification of
specific alleles, or DNA sequence varients
at same locus; specificity is achieved by
designing one or both PCR primers so that
they partially overlap site of sequence
difference between amplified alleles.
Alternative splicing:
Refers to fact that certain genes retain or
omit particular exons in the final spliced
transcript.
Alu sequence: A
family of sequence-related elements about
300 bp in length approximately 500 000
copies of which are scattered along the
human genome.
Amino acid: Any of a
class of 20 molecules that are combined to
form proteins in living things. Consisting
of the basic formula NH2-CHR-COOH, where
"R" is the side chain which
defines the amino acid:The sequence of amino
acids in a protein and hence protein
function are determined by the genetic code.
Nonpolar side chains
(hydrophobic)
G Gly Glycine
A Ala Alanine
V Val Valine
I Ile Isoleucine
L Leu Leucine
F Phe Phenylalanine
P Pro Proline
M Met Methionine
W Trp Tryptophan
C Cys Cysteine
Noncharged polar side
chains (hydrophilic)
S Ser Serine
T Thr Threonine
Y Tyr Tyrosine
N Asn Asparagine
Q Gln Glutamine
Acidic side chains (very
polar, hydrophilic)
D Asp Aspartic Acid
E Glu Glutamic Acid
Basic side chains (very
polar, hydrophilic)
K Lys Lysine
R Arg Arginine
H His Histidine
Amino Terminus:
Refers to the NH2 end of a peptide chain (by
custom drawn at the left of a protein
sequence)
Amp resistance: See
"Antibiotic resistance".
Amplification: An
increase in the number of copies of a
specific DNA fragment; can either be in
vivo or in vitro. See cloning,
polymerase chain reaction.
Anchor Sequence: A
hydrophobic amino acid sequence which fixes
a segment of a newly synthesized,
translocating protein within the lipid
bilayer membrane of the endoplasmic
reticulum.
Aneuploid: A cell
containing a number of chromosomes that is
not an even multiple of the haploid number
(n).
Anneal: The act of
two nucleic acid sequences hydrogen bonding
through complementarity of the bases
determining their sequences. Generally
synonymous with "hybridize".
Antibiotic resistance:
Resistance conferred to the host the ability
to survive a given antibiotic by plasmids
containing resistance genes. If such a
plasmid is present in a host, that host will
not be killed by (moderate levels of)
ampicillin or tetracycline. By transforming
plasmids containing antibiotic resistance
genes, one can get rid of all the
contaminating bacteria which does not have
such plasmid, thus ensuring that the plasmid
will be propagated as the surviving cells
divide.
Antibody: An
immunoglobulin protein produced by
B-lymphocytes of the immune system that
binds to a specific antigen molecule. (See
monoclonal antibodies, polyclonal
antibodies.)
Anticodon: The 3-base
sequence of a tRNA that base-pairs with the
mRNA codon to effect translation of the mRNA
into polypeptide sequence. (See transfer RNA
and messanger RNA).
Antigen: Any foreign
substance, such as a virus, bacterium, or
protein that elicits an immune response by
stimulating the production of antibodies.
Antigenic variation:
Mechanism to ensure rapid sequence variation
of the gene(s) encoding homologues of an
individual protein antigen; usually
involving multiple, related gene copies.
Antisense construct:
Plasmid that contains coding region of gene
placed adjacent to promoter in such
orientation as to cause gene to be
transcribed from DNA strand complementary to
that from which RNA is normally transcribed.
Anti-sense strand:
See Sense strand.
AP-1 site: The
binding site on DNA at which the
transcription "factor" AP-1 binds,
thereby altering the rate of transcription
for the adjacent gene. AP-1 is actually a
complex between c-fos protein and c-jun
protein, or sometimes is just c-jun dimers.
The AP-1 site consensus sequence is (C/G)
TGACT(C/A) A. Also known as the TPA-response
element (TRE). (TPA is a phorbol ester,
tetradecanoyl phorbol acetate, which is a
chemical tumor promoter).
Arrayed library:
Individual primary recombinant clones
(hosted in phage, cosmid, YAC, or other
vector) that are placed in two- dimensional
arrays in microtiter dishes. Each primary
clone can be identified by the identity of
the plate and the clone location (row and
column) on that plate. Arrayed libraries of
clones can be used for many applications,
including screening for a specific gene or
genomic region of interest as well as for
physical mapping. Information gathered on
individual clones from various genetic
linkage and physical map analyses is entered
into a relational database and used to
construct physical and genetic linkage maps
simultaneously; clone identifiers serve to
interrelate the multilevel maps. Compare
library, genomic library.
ATG or AUG: The codon
for methionine; the translation initiation
codon. Usually, protein translation can only
start at a methionine codon (although this
codon may be found elsewhere within the
protein sequence as well). In eukaryotic
DNA, the sequence is ATG; in RNA it is AUG.
Usually, the first AUG in the mRNA is the
point at which translation starts, and an
open reading frame follows - i.e. the
nucleotides taken three at a time will code
for the amino acids of the protein, and a
stop codon will be found only when the
protein coding region is complete.
Attenuation: A form
of gene regulation wherein termination of
transcription is controlled to regulate
overall levels of gene expression.
Avidin: A
glycoprotein which binds to biotin with very
high affinity (Kd = 10-15).
Autoradiography: A
process to detect radioactively labeled
molecules (which usually have been separated
in an SDS-PAGE or agarose gel) based on
their ability to create an image on
photographic or X-ray film. This process
does not result in a linear relationship
between the intensity of the signal and the
amount of radioactivity unless special steps
are taken. There is now increasing use of
phosphorimagers and other modern devices to
detect and quantitate radioactive molecules
which have been separated in gels.
Autosome: A
chromosome not involved in sex
determination. The diploid human genome
consists of 46 chromosomes, 22 pairs of
autosomes, and 1 pair of sex chromosomes
(the X and Y chromosomes).
B
Back Mutation:
Reverse the effect of a point or frame-shift
mutation that had altered a gene; thus it
restores the wild-type phenotype (see
Revertant).
Bacterial artificial
chromosome (BAC): A chromosome-like
structure, constructed by genetic
engineering. BAC is a cloning vector capable
of carrying between 100 and 300 kilobases of
target sequence. They are propagated as a
mini-chromosome in a bacterial host. The
size of the typical BAC is ideal for use as
an intermediate in large-scale genome
sequencing projects. Entire genomes can be
cloned into BAC libraries, and entire BAC
clones can be shotgun-sequenced fairly
rapidly.
Bacteriophage:
(simply phage) A virus that infects a
bacterium and which is often used in
molecular genetics experiments as a vector,
or cloning vehicle. Recombinant phages can
be made in which certain non-essential l DNA
is removed and replaced with the DNA of
interest. The phage can accommodate a DNA
"insert" of about 15-20 kb.
Replication of that virus will thus
replicate the investigator's DNA. One would
use phage l rather than a plasmid if the
desired piece of DNA is rather large.
Bacteriophage l and M13 are ones commonly
used in cloning and/or subcloning of small
genes or DNA fragments in E. coli.
Bacteriophage P1 is one that is used for
fragments up to 95 Kb in size.
Band: A pattern of
light and dark regions by Giemsa staining
that can serve as landmarks on chromosomes.
Band shift assay: see
Gel shift assay.
Base: In molecular
biology, this term refers to the purine
bases adenine and guanine, and the
pyrimidine bases uracil, thymine, and
cytosine, or modification of these bases.
Base Pair (bp): One
pair of complementary nucleotides within a
duplex strand of a nucleic acid. Under
Watson-Crick rules, these pairs consist of
one pyrimidine and one purine: i.e., C-G,
A-T (DNA) or A-U (RNA). However,
"noncanonical" base pairs (e.g.,
G-U) are common in RNA secondary structure.
Base sequence: The
order of nucleotide bases in a DNA molecule.
Base sequence analysis:
A method, sometimes automated, for
determining the base sequence.
B-DNA: Conformation
of the Watson-Crick double helix in which
the two strands form a right-handed double
helix.
Binding site: A place
on cellular DNA to which a protein (such as
a transcription factor) can bind. Typically,
binding sites might be found in the vicinity
of genes, and would be involved in
activating transcription of that gene
(promoter elements), in enhancing the
transcription of that gene (enhancer
elements), or in reducing the transcription
of that gene (silencers). NOTE that whether
the protein in fact performs these functions
may depend on some condition, such as the
presence of a hormone, or the tissue in
which the gene is being examined. Binding
sites could also be involved in the
regulation of chromosome structure or of DNA
replication.
Biotechnology: A set
of biological techniques developed through
basic research and now applied to research
and product development. In particular, the
use by industry of recombinant DNA, cell
fusion, and new bioprocessing techniques.
Biotin: A coenzyme
which is essential for carboxylation
reactions (see Avidin).
Blotting: A technique
for detecting one RNA within a mixture of
RNAs (a Northern blot) or one type of DNA
within of a mixture of DNAs (a Southern
blot). A blot can prove whether that one
species of RNA or DNA is present, how much
is there, and its approximate size.
Basically, blotting involves gel
electrophoresis, transfer to a blotting
membrane (typically nitrocellulose or
activated nylon), and incubating with a
radioactive probe. Exposing the membrane to
X-ray film produces darkening at a spot
correlating with the position of the DNA or
RNA of interest. The darker the spot, the
more nucleic acid was present there.
Blunt End: A terminus
of a duplex DNA molecule which ends
precisely at a base pair, with no overhang
(unpaired nucleotide) in either strand. Some
but not all restriction endonucleases leave
blunt ends after cleaving DNA. Blunt-ended
DNA can be ligated nonspecifically to other
blunt-ended DNA molecules (compare with
Sticky End).
5'-->3'
NNNCCC GGGNNN SmaI cut,
no overhang
NNNGGG CCCNNN
3'<--5'
bp: See base pair.
Box: Refers to a
short nucleic acid consensus sequence or
motif that is universal within kingdoms of
organisms. Examples of DNA boxes are the
Pribow box (TATAAT) for RNA polymerase, the
Hogness box (TATA) that has a similar
function in eukaryotic organisms, and the
homeo box. RNA boxes have also been
described, such as Pilipenko's Box-A motif
that may be involved in ribosome binding in
some viral RNAs.
C
C Terminus:
See Carboxyl Terminus.
Cancer: A disease
process initiated by transformation of a
cell to behavior lacking in normal
proliferative control, and often involving
invasive behavior and localized
vascularization.
Cap: All eukaryotes
have at the 5' end of their messages a
structure called a "cap",
consisting of a 7-methylguanosine in 5'-5'
triphosphate linkage with the first
nucleotide of the mRNA. It is added
post-transcriptionally, and is not encoded
in the DNA.
Cap site: Two usages:
1] In eukaryotes, the cap
site is the position in the gene at which
transcription starts, and really should be
called the "transcription initiation
site". The first nucleotide is
transcribed from this site to start the
nascent RNA chain. That nucleotide becomes
the 5' end of the chain, and thus the
nucleotide to which the cap structure is
attached (see "Cap").
2] In bacteria, the CAP
site (note the capital letters) is a site on
the DNA to which a protein factor (the
Catabolite Activated Protein) binds.
Carboxyl Terminus:
Refers to the COOH end of a peptide chain
(by custom drawn at the right of a protein
sequence)
CAT assay: An enzyme
assay. CAT stands for chloramphenicol acetyl
transferase, a bacterial enzyme which
inactivates chloramphenicol by acetylating
it. CAT assays are often performed to test
the function of a promoter. The gene coding
for CAT is linked onto a promoter
(transcription control region) from another
gene, and the construct is
"transfected" into cultured cells.
The amount of CAT enzyme produced is taken
to indicate the transcriptional activity of
the promoter (relative to other promoters
which must be tested in parallel). It is
easier to perform a CAT assay than it is to
do a Northern blot, so CAT assays were a
common method for testing the effects of
sequence changes on promoter function.
Largely supplanted by the reporter gene
luciferase.
CCAAT box: (CAT box,
CAAT box, and other variants) A sequence
found in the 5' flanking region of certain
genes which is necessary for efficient
expression. A transcription factor
(CCAAT-binding protein, CBP) binds to this
site.
cDNA: See
complementary DNA.
cDNA clone:
"complementary DNA"; a piece of
DNA copied from an mRNA. The term
"clone" indicates that this cDNA
has been spliced into a plasmid or other
vector in order to propagate it. A cDNA
clone may contain DNA copies of such typical
mRNA regions as coding sequence,
5'-untranslated region, 3' untranslated
region or poly(A) tail. No introns will be
present, nor any promoter sequences (or
other 5' or 3' flanking regions). A
"full-length" cDNA clone is one
which contains all of the mRNA sequence from
nucleotide #1 through to the poly(A) tail.
Centimorgan (cM): A
unit of measure of the statistical
probability recombination frequency between
alleles. One centimorgan is equal to a 1%
chance that a marker at one genetic locus
will be separated from a marker at a second
locus due to crossing over in a single
generation. In human beings, 1 centimorgan
is equivalent, on average, to 1 million base
pairs.
Central dogma: A
phrase that refers to the concept of
information flow proceeding only from DNA to
RNA to protein.
Centromere: A
specialized constricted region of a
chromosome to which spindle fibers attach
during cell division at which two sister
chromatids (the two exact copies of each
chromosome that are formed after
replication) are joined, and which attach to
the spindle during cell division.
Chaperone Proteins: A
series of proteins present in the
endoplasmic reticulum which prevent proteins
from folding prematurely and guide the
proper folding of secreted proteins through
a complex series of binding and release
reactions.
Chromatid: A single,
continuous double stranded DNA molecule with
its unique, complete grouping of genetic
information, associated proteins,
higher-order structures, and centromeric and
telomeric regions necessary for separation
and maintenance after replication.
Chromosomes: A
condensed, fibrillar, self- replicating
genetic structures of cells containing the
cellular DNA that bears in its nucleotide
sequence the linear array of genes. In
prokaryotes, chromosomal DNA is circular,
and the entire genome is carried on one
chromosome. Eukaryotic genomes consist of a
number of chromosomes whose DNA is
associated with different kinds of proteins.
Chromosome jumping: A
technique whereby one starts with a piece of
DNA from one region of a chromosome, and
obtains clones from nearby regions without
cloning everything in between (as in
chromosome walking; see below). One round of
jumping yields new clones at distances of
several tens of kb away from the starting
point. In practice, this method is used when
classical genetics proves that a known piece
of DNA is located on the chromosome close to
a gene you would like to clone (such as a
human disease gene). By cloning fragments
some distance away in both directions from
the known fragment, one might obtain (1)
fragments further from the desired gene
(which are discarded); (2) fragments even
more closely linked to the desired gene (in
which case one goes for another round of
jumping); or (3) fragments from within the
desired gene - the optimal result.
Chromosome walking: A
technique for cloning everything in the
genome around a known piece of DNA (the
starting probe). You screen a genomic
library for all clones hybridizing with the
probe, and then figure out which one extends
furthest into the surrounding DNA. The most
distal piece of this most distal clone is
then used as a probe, so that ever more
distal regions can be cloned. This has been
used to move as much as 200 kb away from a
given starting point (an immense
undertaking). Typically used to
"walk" from a starting point
towards some nearby gene in order to clone
that gene. Also used to obtain the remainder
of a gene when you have isolated a part of
it.
Cis: As used in
molecular biology, an interaction between
two sites which are located within the same
molecule. However, a cis-acting protein can
either be one which acts only on the
molecule of DNA from which it was expressed,
or a protein which acts on itself (e.g.,
self-proteolysis).
Cistron: A nucleic
acid segment corresponding to a polypeptide
chain, including the relevant translational
start (initiation) and stop (termination)
codons.
Clones: A group of
cells derived from a single ancestor.
Clone (verb):
Clone (verb):. The
act of duplicating genetic material within a
vector. To clone a piece of DNA, one would
insert it into some type of vector (say, a
plasmid) and put the resultant construct
into a host (usually a bacterium) so that
the plasmid and insert replicate with the
host. An individual bacterium is isolated
and grown and the plasmid containing the
"cloned" DNA is re-isolated from
the bacteria, at which point there will be
many millions of copies of the DNA -
essentially an unlimited supply. Actually,
an investigator wishing to clone some gene
or cDNA rarely has that DNA in a purified
form, so practically speaking, to
"clone" something involves
screening a cDNA or genomic library for the
desired clone. See also "Probe"
for a description of how one might start a
cloning project, and "Screening"
for how the probe in used.
One can also clone more
complex organisms, with considerable
difficulty. The much-publicized Scottish
research that resulted in the sheep 'Dolly'
exemplifies this approach.
Clone (noun): The
term "clone" can refer either to a
bacterium carrying a cloned DNA, or to the
cloned DNA itself.
Clone bank: See
genomic library.
Cloning: to allow it
to be sequenced or studied in some other
way.
Cloning: The process
of generating sufficient copies of a
particular piece of DNA or asexually
producing a group of cells (clones), all
genetically identical, from a single
ancestor. In recombinant DNA technology, the
use of DNA manipulation procedures to
produce multiple copies of a single gene or
segment of DNA is referred to as cloning
DNA.
Cloning vector: DNA
molecule originating from a virus, a
plasmid, or the cell of a higher organism
into which another DNA fragment of
appropriate size can be integrated without
loss of the vectors capacity for self-
replication; vectors introduce foreign DNA
into host cells, where it can be reproduced
in large quantities. Examples are plasmids,
cosmids, bacterial artificial chromosomes
(BAC) and yeast artificial chromosomes
(YAC); vectors are often recombinant
molecules containing DNA sequences from
several sources.
cM: See centimorgan.
Code: See codon and
genetic code.
Coding sequence: The
portion of a gene or an mRNA which actually
codes for a protein. Introns are not coding
sequences; nor are the 5' or 3' untranslated
regions (or the flanking regions, for that
matter - they are not even transcribed into
mRNA). The coding sequence in a cDNA or
mature mRNA includes everything from the AUG
(or ATG) initiation codon through to the
stop codon, inclusive.
Coding strand: an
ambiguous term intended to refer to one
specific strand in a double-stranded gene.
See "Sense strand".
Codon: A group of
three consecutive nucleotides within
messenger RNA (mRNA) that encodes a message
to initiate translation, to incorporate a
specific amino acid into the growing
polypeptide chain, or to stop translation.
The sequence of codons in the mRNA
unambiguously defines the primary structure
of the final protein. Of course, the codons
in the mRNA were also present in the genomic
DNA, but the sequence may be interrupted by
introns. (See genetic code also).
Codon Bias: The
tendency for an organism or virus to use
certain codons more than others to encode a
particular amino acid. An important
detrminant of codon bias is the
guanosine-cytosine (GC) content of the
genome. An organism that has a relatively
low G+C content of 30% will be less likely
to have a G or C at the third position of a
codon (wobble position) than a A or T to
specify an amino acid that can be
represented by more than one codon.
Co-linearity: Refers
to an exact correspondence between
information encoded in DNA and the
polypeptide product ultimately translated
from transcripts made from the DNA. Most
prokaryotic genes are co-linear with their
products; eukaryotic genes often are not.
Competent: Bacterial
cells which are capable of accepting foreign
extra-chromosomal DNA. There are a variety
of processes by which cells may be made
competent.
Complement:
Definition determined by context:
1] The complementary
sequence to a nucleic acid sequence under
study.
2] To provide a function
that is missing or altered because of a
mutational event.
Complementary DNA (cDNA):
A DNA sequence synthesized from a messenger
RNA molecule, using reverse transcriptase
enzyme. cDNAs can be used experimentally to
determine the sequence of messenger RNAs
after their introns (non-protein-coding
sections) have been spliced out. The single-
stranded form is often used as a probe in
physical mapping.
Complementary sequences:
Nucleic acid base sequences that can form a
double- stranded structure by matching base
pairs; the complementary sequence to G- T-
A- C is C- A- T- G.
Concatemer: Tandem
arrays of monomeric DNA molecules with
complementary ends. Intra-molecular
reassociation of such molecules leads to
circularisation while inter-molecular
reaction produces concatemers.
Conformation: The
3-dimensional structure of a molecule.
Conformational epitope:
Also called a "global epitope". An
epitope comprised of contiguous but
physically discontinuous components of the
immunogenic molecule. In a proteinaceous
antigen this could be sequences from
different stretches within a polypeptide, or
even from different polypeptide subunits.
Conjugation: Physical
contact with the establishment of plasma
bridges between two different bacterial
cells allowing directed transfer of DNA.
Consensus sequence: A
term that refers to sequences common to
different genes within an organism, or to
the same gene among different organisms,
that encode a specific function. This term
may be applied to either nucleic acids or
proteins, since the protein sequence is
completely dependent upon the nucleic acid
sequence.
Conservation:
Identical parts of genes that are present in
two distinct organisms are said to be
conserved. Conservation can be detected by
measuring the similarity of the two
sequences at the base (RNA or DNA) or
amino-acid (protein) level.
Conserved sequence: A
base sequence in a DNA molecule (or an amino
acid sequence in a protein) that has
remained essentially unchanged throughout
evolution. The more similarities there are,
the more highly conserved the two sequences.
Consensus sequence: A
'nominal' sequence inferred from multiple,
imperfect examples. Multiple lanes of
shotgun sequence can me merged to show a
consensus sequence. The optimal sequence of
nucleotides recognized by some factor. A DNA
binding site for a protein may vary
substantially, but one can infer the
consensus sequence for the binding site by
comparing numerous examples. For example,
the (fictitious) transcription factor ZQ1
usually binds to the sequences AAAGTT,
AAGGTT or AAGATT. The consensus sequence for
that factor is said to be AARRTT (where R is
any purine, i.e. A or G). ZQ1 may also be
able to weakly bind to ACAGTT (which differs
by one base from the consensus).
Conservative
Substitution: A nucleotide mutation
which alters the amino acid sequence of the
protein, but which causes the substitution
of one amino acid with another which has a
side chain with similar charge/polarity
characteristics (see Amino Acid). The size
of the side chain may also be an important
consideration. Conservative mutations are
generally considered unlikely to profoundly
alter the structure or function of a
protein, but there are many exceptions (see
Nonconservative Substitution).
Constitutive expression:
Constantly expressed at some appreciable
level.
Contig: Several uses,
all nouns. The term comes from a shortening
of the word 'contiguous'. A 'contig' may
refer to a map showing placement of a set of
clones that completely, contiguously cover
some segment of DNA in which you are
interested. Also called the 'minimal tiling
path'. More often, the term 'contig' is used
to refer to the final product of a shotgun
sequencing project. When individual lanes of
sequence information are merged to infer the
sequence of the larger DNA piece, the
product consensus sequence is called a
'contig'.
Contig map: A map
depicting the relative order of a linked
library of small overlapping clones
representing a complete chromosomal segment.
Cosmid: A type of
artificially constructed vector used for
cloning 35-45 kb of DNA. These are plasmids
carrying a phage l cos site (which allows
packaging into l capsids), an origin of
replication and an antibiotic resistance
gene. A plasmid of 40 kb is very difficult
to put into bacteria, but can replicate once
there. Cosmids, however, have a cos site,
and thus can be packaged into l phage heads
(a reaction which can be performed in
vitro) to allow efficient introduction
into bacteria.
Cos site:
Crossing over: The
breaking during meiosis of one maternal and
one paternal chromosome, the exchange of
corresponding sections of DNA, and the
rejoining of the chromosomes. This process
can result in an exchange of alleles between
chromosomes. Compare recombination.
Cytosine (C): One of
the pyrimidine bases found in DNA and RNA
(4-amino-2-hydroxypyrimidine). one member of
the base pair G- C (guanine and cytosine).
D
Dalton:
Unit
of atomic mass. One dalton corresponds to
the mass of a hydrogen atom = 3.32X10-24
g.
Database Search: Once
an open reading frame or a partial amino
acid sequence has been determined, the
investigator compares the sequence with
others in the databases using a computer and
a search algorithm. This is usually done in
a protein database such as PIR or
Swiss-Prot. Nucleic acid sequences are in
GenBank and EMBL databases. The search
algorithms most commonly used are BLAST and
FASTA.
Degeneracy: In
molecular biology, this term refers to the
fact that multiple different codons may
encode the same amino acid. However, a given
codon does not encode more than one amino
acid within the nucleus of an organism.
Denaturation: With
respect to nucleic acids, refers to the
conversion from double-stranded to the
single-stranded state, often achieved by
heating or alkaline conditions. This is also
called "melting" DNA. With respect
to proteins, refers to the disruption of
tertiary and secondary structure, often
achieved by heat, detergents, chaotropes,
and sulfhydryl-reducing agents.
Denaturing Gel: An
agarose or acrylamide gel run under
conditions which destroy secondary or
tertiary protein or RNA structure. For
protein, this usually means the inclusion of
2-ME (which reduces disulfide bonds between
cysteine residues) and SDS and/or urea in an
acrylamide gel. For RNA, this usually means
the inclusion of formaldehyde or glyoxal to
destroy higher ordered RNA structures. In
DNA sequencing gels, urea is included to
denature dsDNA to ssDNA strands. In
denaturing gels, macromolecules tend to be
separated on the basis of size and (to some
extent) charge, while shape and
oligomerization of molecules are not
important. Contrast with Native Gel.
Denaturing gradient gel
electrophoresis (DGGE): Resolves partially
denatured double stranded DNA in precisely
conditions of temperature and denaturant
concentration. Different alleles may
denature to various extents under such
conditions, and migrate differently on DGGE
acrylamide gels.
Deletion: The absence
of bases that are present in the wild-type
DNA sequence.
Deoxyribonucleotide:
See nucleotide.
Dideoxy Sequencing:
Enzymatic determination of DNA or RNA
sequence by the method of Sanger and
colleagues, based on the incorporation of
chain terminating dideoxynucleotides in a
growing nucleic acid strand copied by DNA
polymerase or reverse transcriptase from a
DNA or RNA template. Separate reactions
include dideoxynucleotides containing A,C,
G, or T bases. The reaction products
represent a collection of new. labeled DNA
strands of varying lengths, all terminating
with a dideoxynucleotide at the 3' end (at
the site of a complementary base in the
template nucleic acid), and are separated in
a polyacrylamide/urea gel to generate a
sequence "ladder". This method is
more commonly used than
"Maxam-Gilbert" (chemical)
sequencing.
Direct Repeats:
Identical or related sequences present in
two or more copies in the same orientation
in the same molecule of DNA; they are not
necessarily adjacent.
Diploid: A full set
of genetic material, consisting of paired
chromosomes one chromosome from each
parental set. Most animal cells except the
gametes have a diploid set of chromosomes.
The diploid human genome has 46 chromosomes.
Compare haploid.
DNA (deoxyribonucleic
acid): The molecule responsible for
storing and transmitting genetic
information. DNA is a double- stranded
molecule held together by weak bonds between
base pairs of nucleotides twisted around
each other in the shape of a double helix.
The four nucleotides in DNA contain the
bases: adenine (A), guanine (G), cytosine
(C), and thymine (T). In nature, base pairs
form only between A and T and between G and
C; thus the base sequence of each single
strand can be deduced from that of its
partner.
DNA gyrase: see
gyrase.
DNA ligase: Enzymatic
activity responsible for creating
phosphodiester bonds between the 5' end of
one strand of DNA and the 3' end of another
strand. Requires the presence of a 5'
phosphate on one strand, and a 3' Hydroxyl
group on the second strand.
DNA polymerase:
Enzymatic activity responsible for
catalyzing the polymerization of DNA. Is
dependent upon an annealed primer from which
to initiate polymerization, and a DNA
template from which to copy.
DNA probes: See
probe.
DNA replication: The
use of existing DNA as a template for the
synthesis of new DNA strands. In humans and
other eukaryotes, replication occurs in the
cell nucleus.
DNase:
Deoxyribonuclease, a class of enzymes which
digest DNA. The most common is DNase I, an
endonuclease which digests both single and
double-stranded DNA.
DNA sequence: The
relative order of base pairs, whether in a
fragment of DNA, a gene, a chromosome, or an
entire genome. See base sequence analysis.
Domain: A discrete
portion of a protein with its own function.
The combination of domains in a single
protein determines its overall function.
Dominant: Allele that
determines phenotype in a heterozygous
individual carrying another recessive
allele.
Dot blot: A technique
for measuring the amount of one specific DNA
or RNA in a complex mixture. The samples are
spotted onto a hybridization membrane (such
as nitrocellulose or activated nylon, etc.),
fixed and hybridized with a radioactive
probe. The extent of labeling (as determined
by autoradiography and densitometry) is
proportional to the concentration of the
target molecule in the sample. Standards
provide a means of calibrating the results.
Double helix: The
helical shape assumed by DNA in which the
two complementary strands hydrogen bond
together in opposite orientations (i.e. have
opposite polarities).
Downstream: See
"Upstream/Downstream".
Duplex: A nucleic
acid molecule in which two strands are base
paired with each other.
E
E. coli (Escherichia
coli):
A common Gram-negative bacterium present in
human intestinal tract that has been studied
intensively by geneticists because of its
small genome size, normal lack of
pathogenicity, and ease of growth in the
laboratory. Useful for cloning experiments.
Electroporation: A
method for introducing foreign nucleic acid
into bacterial or eukaryotic cells that uses
a brief, high voltage DC charge which
renders the cells permeable to the nucleic
acid. Also useful for introducing synthetic
peptides into eucaryotic cells.
Electrophoresis: A
method of separating large molecules (such
as DNA fragments or proteins) from a mixture
of similar molecules. An electric current is
passed through a medium containing the
mixture, and each kind of molecule travels
through the medium at a different rate,
depending on its electrical charge and size.
Separation is based on these differences.
Agarose and acrylamide gels are the media
commonly used for electrophoresis of
proteins and nucleic acids.
Elongation factor: A
protein(s) that associates with the ribosome
cyclically to assist in loading tRNA into
the the A site of the ribosome.
End Labeling: The
technique of adding a radioactively labeled
group to one end (5' or 3' end) of a DNA
strand.
Endonuclease: An
enzyme that cleaves its nucleic acid
substrate at internal sites (as opposed to
an exonuclease, which must start at an end)
in the nucleotide sequence. Examples include
the restriction enzymes, DNase I and RNase
A.
Endoplasmic reticulum:
A specialized membranous organelle within
eukaryotic cells responsible for synthesis
of membrane-inserted proteins, and for
proteins to be exported of proteins to the
cell surface or beyond.
Enhancer: An enhancer
is a cis-acting nucleotide sequence to which
transcription factor(s) bind, and which
increases the transcription of a gene. It is
not part of a promoter; the basic difference
being that an enhancer can be moved around
anywhere in the general vicinity of the gene
(within several thousand nucleotides on
either side or even within an intron), and
it will still function. It can even be
clipped out and spliced back in backwards,
and will still operate. A promoter, on the
other hand, is position- and
orientation-dependent. Enhancers are ususlly
around 70-80 bp in length and are found, for
e.g in viral DNA molecules. Some enhancers
are "conditional" - in other
words, they enhance transcription only under
certain conditions, for example in the
presence of a hormone.
Enzyme: A protein
that acts as a catalyst, speeding the rate
at which a biochemical reaction proceeds but
not altering the direction or nature of the
reaction.
Epigenetic: A change
in phenotype brought about by changes in
gene regulation rather than by a change in
genotype.
Episome:
Extrachromosomal genetic element. Generally
used synonymously for plasmid.
Epitope: As related
to protein antigens, B-cell epitopes consist
of the amino acid residues of a protein
molecule which interact directly through
noncovalent bonds with the amino acid
residues of a particular antibody molecule
(complementarity determining region). The
average epitope probably involves about
15-20 contact amino acid residues, but one
or two of these may be critical to the
epitope's specificity and the avidity of the
antibody-antigen reaction. B-cell epitopes
may be either linear or conformational in
nature. T-cell epitopes represent the small,
processed peptides which bind to MHC class I
and II molecules on the surface of T cells.
EST: see expressed
sequence tag and sequence tagged site (STS).
ERE: Estrogen
Response Element. A binding site in a
promoter to which the activated estrogen
receptor can bind. The estrogen receptor is
essentially a transcription factor which is
activated only in the presence of estrogens.
The activated receptor will bind to an ERE,
and transcription of the adjacent gene will
be altered. See also "Response
element".
Ethidium Bromide:
Intercalates within the structure of nucleic
acids in such a way that they fluoresce
under UV light. Ethidium bromide staining is
commonly used to visualize RNA or DNA in
agarose gels placed on UV light boxes.
Proper precautions are required, because the
ethidium bromide is highly mutagenic and the
UV light damaging to the eyes. Ethidium
bromide is also included in cesium chloride
gradients during ultracentrifugation, to
separate supercoiled circular DNA from
linear and relaxed circular DNA.
Euchromatin: The
gene-rich regions of a genome (see also
heterochromatin).
Eukaryote: Cell or
organism with membrane- bound, structurally
discrete nucleus and other well- developed
subcellular compartments. Eukaryotes include
all organisms except viruses, bacteria, and
blue- green algae. Compare prokaryote. See
chromosomes.
Evolutionary Clock:
Defined by the rate at which mutations
accumulate within a given gene.
Evolutionarily conserved:
See conserved sequence.
Exogenous DNA: DNA
originating outside an organism.
Exon: Those portions
of a genomic DNA sequence which will be
represented in the final, mature mRNA ie. A
contiguous segment of genomic DNA that codes
for a polypeptide in a gene.The term
"exon" can also be used for the
equivalent segments in the final RNA. Exons
may include coding sequences, the 5'
untranslated region or the 3' untranslated
region.
Exonuclease: An
enzyme that cleaves nucleotides sequentially
from free ends of a linear nucleic acid
substrate. An example is Exonuclease III,
which digests only double-stranded DNA
starting from the 3' end.
Expressed sequence tag
(EST): A short piece of DNA sequence
corresponding to a fragment of a
complementary DNA (made from a cell's
messenger RNA). ESTs have been used to hunt
for genes, so hundreds of thousands are
present in sequence databases. (See Sequence
tagged sites)
Expression: To
"express" a gene is to cause it to
function. A gene which encodes a protein
will, when expressed, be transcribed and
translated to produce that protein. A gene
which encodes an RNA rather than a protein
(for example, a rRNA gene) will produce that
RNA when expressed.
Expression clone:
This is a clone (plasmid in a bacteria, or
maybe a lambda phage in bacteria) which is
designed to produce a protein from the DNA
insert. Mammalian genes do not function in
bacteria, so to get bacterial expression
from your mammalian cDNA, you would place
its coding region (i.e. no introns)
immediately adjacent to bacterial
transcription/translation
control sequences. That artificial construct
(the "expression clone") will
produce a pseudo-mammalian protein if put
back into bacteria. Often, that protein can
be recognized by antibodies raised against
the authentic mammalian protein, and vice
versa.
Expression Vector: A
plasmid or phage designed for production of
a polypeptide from inserted foreign DNA
under specific controls. Often an inducer is
used. The vector always provides a promoter
and often the transcriptional start site,
ribosomal binding sequence, and initiation
codon. In some cases the product is a fusion
protein.
Expressed gene: See
gene expression.
F
Familial:
An inherited trait.
FISH (fluorescence in
situ hybridization): A physical mapping
approach that uses fluorescein tags to
detect hybridization of probes with
metaphase chromosomes and with the less-
condensed somatic interphase chromatin.
Flow cytometry:
Analysis of biological material by detection
of the light- absorbing or fluorescing
properties of cells or subcellular fractions
(i.e., chromosomes) passing in a narrow
stream through a laser beam. An absorbance
or fluorescence profile of the sample is
produced. Automated sorting devices, used to
fractionate samples, sort successive
droplets of the analyzed stream into
different fractions depending on the
fluorescence emitted by each droplet.
Flow karyotyping: Use
of flow cytometry to analyze and/or separate
chromosomes on the basis of their DNA
content.
Footprinting: A
technique by which one identifies a protein
binding site on cellular DNA. The presence
of a bound protein prevents DNase from
"nicking" that region, which can
be detected by an appropriately designed
gel.
Frame-shift: A change
from one reading frame to another.
Frameshift Mutation:
A mutation (deletion or insertion, never a
simple substitution) of one or more
nucleotides but never a multiple of 3
nucleotides, which shortens or lengthens a
trinucleotide sequence representing a codon;
the result is a shift from one reading frame
to another reading frame. The amino acid
sequence of the protein downstream of the
mutation is completely altered, and may even
be much shorter or longer due to a change in
the location of the first termination (stop)
codon:
Asn Tyr Thr Asn Leu Gly
His Wild-type polypeptide
AAU UAC ACA AAU UUA GGG
CAU mRNA
Asn Thr Gln Ile STOP
Mutant polypeptide
|
Deletion of A from mRNA
creates frame-shift mutant
Fusion Protein: A
product of recombinant DNA in which the
foreign gene product is juxtaposed
("fused") to either the
carboxyl-terminal or amino-terminal portion
of a polypeptide encoded by the vector
itself. Use of fusion proteins often
facilitates expression of otherwise lethal
products and the purification of recombinant
proteins.
G
gDNA: Shorthand for
"genomic DNA".
Gel shift assay: (gel
mobility shift assay, band shift assay) A
method by which one can determine whether a
particular protein preparation contains
factors which bind to a particular DNA
fragment. When a radiolabeled DNA fragment
is run on a gel, it shows a characteristic
mobility. If it is first incubated with a
cellular extract of proteins (or with
purified protein), any protein-DNA complexes
will migrate slower than the naked DNA - a
shifted band.
Gamete: Mature male
or female reproductive cell (sperm or ovum)
with a haploid set of chromosomes (23 for
humans).
Gene: The fundamental
physical and functional unit of heredity
(Usually DNA, Some organisms have RNA a
gene). A gene is an ordered sequence of
nucleotides located in a particular position
on a particular chromosome that encodes a
specific functional product (i.e., a protein
or RNA molecule) that contributes to or
influences the phenotype of the cell. A gene
usually contains coding regions, introns,
untranslated regions and control regions.
See gene expression.
Gene Conversion: The
alteration of all or part of a gene by a
homologous donor DNA that is itself not
altered in the process.
Gene expression: The
process by which information coded by genes
is converted into the structures present and
operating in the cell. Expressed genes
include those that are transcribed into mRNA
and then translated into protein and those
that are transcribed into RNA but not
translated into protein (e.g., transfer and
ribosomal RNAs).
Gene families: Groups
of closely related genes that make similar
products.
Gene library: See
genomic library.
Gene mapping:
Determination of the relative positions of
genes on a DNA molecule (chromosome or
plasmid) and of the distance, in linkage
units or physical units, between them.
Gene product: The
biochemical material, either RNA or protein,
resulting from expression of a gene. The
amount of gene product is used to measure
how active a gene is; abnormal amounts can
be correlated with disease- causing alleles.
Gene superfamily: A
large group of genes related (often poorly)
by sequence homologies or by the structures
of their products, and by their involvement
in different aspects of the same larger
process (e.g. immunoglobulin gene
superfamily).
Gene therapy: A
technique involving the use of foreign
genetic material to correct a genetic defect
or to modify the phenotype of an affected
individual, by targeting the somatic cells.
Genetic code: The
sequence of nucleotides, coded in triplets
(codons) along the mRNA, that determines the
sequence of amino acids in protein
synthesis. The DNA sequence of a gene can be
used to predict the mRNA sequence, and the
genetic code can in turn be used to predict
the amino acid sequence.
Genetic disease:
Heritable genetic alterations from wild type
which, when expressed, result in decreased
viability of the individual receiving the
altered gene(s).
Genetic engineering
technologies: See recombinant DNA
technologies.
Genetic map: The
ordering of genes by the statistical
determination of recombination events
between them. Genes separated by greater
distances are more likely to recombine. (See
also linkage map).
Genetic material: See
genome.
Genetics: The study
of the patterns of inheritance of specific
traits.
Genome: The entire
compliment of genetic material in the form
of permanently maintained DNA for a given
organism. Its size is generally given as its
total number of base pairs. Mammalian
genomic DNA (including that of humans)
contains 6 billion base pairs of DNA per
diploid cell. There are somewhere in the
order of a hundred thousand genes, including
coding regions, 5' and 3' untranslated
regions, introns, 5' and 3' flanking DNA.
Also present in the genome are structural
segments such as telomeric and centromeric
DNAs and replication origins, and intergenic
DNA.
Genome projects:
Research and technology development efforts
aimed at mapping and sequencing some or all
of the genome of human beings and other
organisms.
Genomic blot: A type
of Southern blot specifically used to
analyze a mixture of DNA fragments derived
from total genomic DNA. Because genomic DNA
is very complicated, when it has been
digested with restriction enzymes, it
produces a complex set of fragments ranging
from tens of bp to tens of thousands of bp.
However, any specific gene will be
reproducibly found on only one or a few
specific fragments. A million identical
cells will produce a million identical
restriction fragments for any given gene, so
probing a genomic Southern with a
gene-specific probe will produce a pattern
of perhaps one or just a few bands.
Genomic clone: A
piece of DNA taken from the genome of a cell
or animal, and spliced into a bacteriophage
or other cloning vector. A genomic clone may
contain coding regions, exons, introns, 5'
flanking regions, 5' untranslated regions,
3' flanking regions, 3' untranslated
regions, or it may contain none of
these...it may only contain intergenic DNA
(usually not a desired outcome of a cloning
experiment!).
Genomic library: A
collection of clones made from a set of
randomly generated overlapping DNA fragments
representing the entire genome of an
organism. Compare library, arrayed library.
Genotype: The set of
genes that an individual carries; usually
refers to the particular pair of alleles
(alternative forms of a gene) that a person
has at a given region of the genome.
Germline: Tissues
involved in the generation of haploid
gametes.
Glycosylation: The
covalent addition of sugar moities to N or O
atoms present in the side chains of certain
amino acids of certain proteins, generally
occuring within the Golgi apparatus during
secretion of a protein. Glycosylation sites
are only partially predictable by current
computer searches for relevant motifs in
protein sequence. Glycosylation may have
profound but very unpredictable effects on
the folding, stability, and antigenicity of
secreted proteins. Glycosylation is a
property of eukaryotic cells, and differs
among different cell types (i.e., it may be
very different in yeast or insect cells used
for protein expression, when compared with
Chinese hamster ovary (CHO) cells).
Golgi Apparatus: A
membranous, vesicular structure which is in
continuity with the endoplasmic reticulum of
eukaryotic cells and generally in close
proximity to the nucleus, the Golgi plays an
important role in the posttranslational
processing and transport of secreted
proteins.
GRE: Glucocorticoid
Response Element: A binding site in a
promoter to which the activated
glucocorticoid receptor can bind. The
glucocorticoid receptor is essentially a
transcription factor which is activated only
in the presence of glucocorticoids. The
activated receptor will bind to a GRE, and
transcription of the adjacent gene will be
altered. See also "Response
element".
Guanine (G): A
nitrogenous base found in DNA and RNA, one
member of the base pair G- C (guanine and
cytosine). (2-amino-6-hydroxypurine).
Gyrase: An enzymatic
activity responsible for maintaining
supercoiling in DNA.
H
Hairpin:
A helical (duplex) region formed by base
pairing between adjacent (inverted)
complementary sequences within a single
strand of RNA or DNA.
Haploid: A single set
of chromosomes (half the full set of genetic
material), present in the egg and sperm
cells of animals and in the egg and pollen
cells of plants. Human beings have 23
chromosomes in their reproductive cells.
Compare diploid.
Haplotype: A
particular combination of alleles
(alternative forms of genes) or sequence
variations that are closely linked that
is, are likely to be inherited together
on the same chromosome. Originally was
coined to describe MHC antigens, but now is
used to describe RFLP patterns and certain
other situations.
H-chain:
Immunoglobulin heavy chain.
Helix-turn-helix: A
protein structural motif characteristic of
certain DNA-binding proteins.
Heteroduplex Dna:
Generated by base pairing between
complementary single strands derived from
different parental duplex molecules;
heteroduplex DNA molecules occur during
genetic recombination in vivo and during
hydridization of different but related DNA
strands in vitro. Since the sequences of the
two strands in a heteroduplex differ, the
molecule is not perfectly base-paired; the
melting temperature of a heteroduplex DNA is
dependent upon the number of mismatched base
pairs.
Heterozygous: An
individual containing dissimilar alleles for
a given gene or locus.
hnRNA (heterogeneous
nuclear RNA): Heterogeneous nuclear RNA;
refers collectively to the variety of RNAs
found in the nucleus, including primary
transcripts, partially processed RNAs and
snRNA. The term hnRNA is often used just for
the unprocessed primary transcripts,
however.
Heterochromatin:
Compact, gene-poor regions of a genome,
which are enriched in simple sequence
repeats. As it can be impossible to clone,
heterochromatin is often ignored when
calculating the percentage of a genome that
has been sequenced. Heterochromatin was
originally identified as regions of the
genome that stained differently to
euchromatin (gene-rich regions).
Heterozygosity: The
presence of different alleles at one or more
loci on homologous chromosomes.
Histones: Highly
basic proteins which associate with the
chromosomal DNA to package it into a
compact, higher order structure.
Homeobox: A short
stretch of nucleotides whose base sequence
is virtually identical in all the genes that
contain it. It has been found in many
organisms from fruit flies to human beings.
In the fruit fly, a homeobox appears to
determine when particular groups of genes
are expressed during development.
Homeotic mutation: A
mutation in a homeotic gene that is
responsible for controlling the activities
of numerous other genes, usually during
embryologic development in higher organisms.
Homologous Recombination:
The exchange of sequence between two related
but different DNA (or RNA) molecules, with
the result that a new "chimeric"
molecule is created. Several mechanisms may
result in recombination, but an essential
requirement is the existence of a region of
homology in the recombination partners. In
DNA recombination, breakage of single
strands of DNA in the two recombination
partners is followed by joining of strands
present in opposing molecules, and may
involve specific enzymes. Recombination of
RNA molecules may occur by other mechanisms.
Homologous chromosomes:
A pair of chromosomes containing the same
linear gene sequences, each derived from one
parent.
Homology: Indicates
similarity between two different nucleotide
or amino acid sequences, often with
potential evolutionary significance. It is
probably better to use more quantitative and
descriptive terms such as nucleotide
"identity" or, in the case of
proteins, amino acid "identity" or
"relatedness" (the latter refers
to the presence of amino acids residues with
similar polarity/charge characteristics at
the same position within a protein).
Homozygous: An
individual containing identical alleles for
a given gene or locus.
Host strain (bacterial):
The bacterium used to harbor a plasmid.
Typical host strains include HB101
(general purpose E. coli strain), DH5a
(ditto), JM101 and JM109
(suitable for growing M13 phages), XL1-Blue
(general-purpose, good for blue/white lacZ
screening). Note that the host strain is
available in a form with no plasmids (hence
you can put one of your own into it), or it
may have plasmids present (especially if you
put them there). Hundreds, perhaps
thousands, of host strains are available.
Human gene therapy:
Insertion of normal DNA directly into cells
to correct a genetic defect.
Human Genome Initiative:
Collective name for several projects begun
in 1986 by Department Of Energy (DOE) to (1)
create an ordered set of DNA segments from
known chromosomal locations, (2) develop new
computational methods for analyzing genetic
map and DNA sequence data, and (3) develop
new techniques and instruments for detecting
and analyzing DNA. This DOE initiative is
now known as the Human Genome Program. The
national effort, led by DOE and NIH, is
known as the Human Genome Project.
Hybridization: The
reaction by which the pairing of
complementary strands of nucleic acid
occurs. DNA is usually double-stranded, and
when the strands are separated they will
re-hybridize under the appropriate
conditions. Hybrids can form between
DNA-DNA, DNA-RNA or RNA-RNA. They can form
between a short strand and a long strand
containing a region complementary to the
short one. Imperfect hybrids can also form,
but the more imperfect they are, the less
stable they will be (and the less likely to
form). To "anneal" two strands is
the same as to "hybridize" them.
Hybridoma: A clone of
plasmacytoma cells which secrete a
monoclonal antibody; usually produced by
fusion of peripheral or splenic plasma cells
taken from an immunized mouse with an
immortalized murine plasmacytoma cell line
(fusion partner), followed by cloning and
selection of appropriate antibody-producing
cells.
Hydrophilicity Plot:
A computer plot which examines the relative
summed hyrophobicity/hydrophilicity of
adjacent amino acid sidechains (usually
within a moving window of about 6 amino acid
residues) along the primary sequence of a
polypeptide chain. Values for the
contribution of sidechains of each the 20
common amino acids to
hydrophobicity/hydrophilicity have been
developed by Hopp & Woods, and Kyte
& Doolittle, and these plots are often
named after these workers. Generally,
hydrophobic regions of proteins are
considered likely to be in the interior of
the native protein, while hydrophilic
domains are likely to be exposed on the
surface and thus possibly antigenic sites
(epitopes). At best, these are crude
predictions.
I
Idiotope:
Epitopes present in the unique variable
sequences of an immunoglobulin molecule.
These may or may not coincide with the
immunoglobulin's paratope.
Immunoprecipitation:
A process whereby a particular protein of
interest is isolated by the addition of a
specific antibody, followed by
centrifugation to pellet the resulting
immune complexes. Often, staphylococcal
proteins A or G, bound to sepharose or some
other type of macroscopic particle, is added
to the reaction mix to increase the size and
ease collection of the complexes. Usually,
the precipitated protein is subsequently
examined by SDS-PAGE.
Inducer: A small
molecule, such as IPTG, that triggers gene
transcription by binding to a regulator
protein, such as LacZ.
Informatics: The
study of the application of computer and
statistical techniques to the management of
information. In genome projects, informatics
includes the development of methods to
search databases quickly, to analyze DNA
sequence information, and to predict protein
sequence and structure from DNA sequence
data.
Initiation Codon: The
codon at which translation of a polypeptide
chain is initiated. This is usually the
first AUG triplet in the mRNA molecule from
the 5' end, where the ribosome binds to the
cap and begins to scan in a 3' direction.
However, the surrounding sequence context is
important and may lead to the first AUG
being bypassed by the scanning ribosome in
favor of an alternative, downstream AUG.
Also called a "start codon".
Occasionally other codons may serve as
initiation codons, e.g. UUG.
Initiator tRNA: A
special tRNA responsible for annealing to
the start codon, in the P site of the
ribosome, to initiate polypeptide synthesis.
Insert: In a complete
plasmid clone, there are two types of DNA -
the "vector" sequences and the
"insert". The vector sequences are
those regions necessary for propagation,
antibiotic resistance, and all those mundane
functions necessary for useful cloning. In
contrast, however, the insert is the piece
of DNA in which you are really interested.
Insertion: The
presence of additional bases within a
sequence that are not present in wild-type
sequence.
Insertion Sequence: A
small bacterial transposon carrying only the
genetic functions involved in transposition.
There are usually inverted repeats at the
ends of the insertion sequence.
In situ
hybridization: Use of a DNA or RNA probe
to detect the presence of the complementary
DNA sequence in cloned bacterial or cultured
eukaryotic cells.
Intergenic: Between
two genes; e.g. intergenic DNA is the DNA
found between two genes. The term is often
used to mean non-functional DNA (or at least
DNA with no known importance to the two
genes flanking it). Alternatively, one might
speak of the "intergenic distance"
between two genes as the number of base
pairs from the polyA site of the first gene
to the cap site of the second. This usage
might therefore include the promoter region
of the second gene.
Interphase: The
period in the cell cycle when DNA is
replicated in the nucleus; followed by
mitosis.
Intron: Introns are
portions of genomic DNA which are
transcribed (and thus present in the primary
transcript) but which are later spliced out.
They thus are not present in the mature
mRNA. Note that although the 3' flanking
region is often transcribed, it is removed
by endonucleolytic cleavage and not by
splicing. It is not an intron. Compare
exons.
Inverse PCR:
Variation of PCR that makes the
amplification of DNA segments of unknown
sequence that flank DNA segments of known
sequence possible; in brief, total DNA is
digested to completion and fragments ligated
under conditions that favour circularization
of fragments; pair of PCR primers, designed
from known sequence known sequence, are used
to prime PCR from opposite strands resulting
in amplification of fragment of unknown
sequence.
Inverted Repeats: Two
copies of the same or related sequence of
DNA repeated in opposite orientation on the
same molecule (contrast with Direct
Repeats). Adjacent inverted repeats
constitute a palindrome.
In vitro:
Literally means "in glass", e.g.,
test tube; now used to mean growth in any
type of culture vessel; Outside a living
organism.
In vivo: In
living organism
K
Karyotype:
A
photomicrograph of an individuals
chromosomes arranged in a standard format
showing the number, size, and shape of each
chromosome type; used in low- resolution
physical mapping to correlate gross
chromosomal abnormalities with the
characteristics of specific diseases.
kb:. abbreviation for
kilobase, one thousand bases. See kilobase
kd: one thousand
Dalton. See Dalton.
Kilobase (kb): Unit
of length for DNA fragments equal to 1000
nucleotides.
Kinase: A kinase is
in general an enzyme that catalyzes the
transfer of a phosphate group from ATP to
something else. In molecular biology, it has
acquired the more specific verbal usage for
the transfer onto DNA of a radiolabeled
phosphate group. This would be done in order
to use the resultant "hot" DNA as
a probe.
Kinteochore: A
specialized structure found in the
centromeric region of the chromosome that is
responsible for attaching to the spindle
during nuclear division.
Klenow Fragment: The
large fragment of E. coli DNA polymerase I
which lacks 5' -> 3' exonuclease
activity. Very useful for sequencing
reactions, which proceed in a 5' -> 3'
fashion (addition of nucleotides to
templated free 3' ends of primers).
Knock-out experiment:
A technique for deleting, mutating or
otherwise inactivating a gene in a mouse.
This laborious method involves transfecting
a crippled gene into cultured embryonic stem
cells, searching through the thousands of
resulting clones for one in which the
crippled gene exactly replaced the normal
one (by homologous recombination), and
inserting that cell back into a mouse
blastocyst. The resulting mouse will be
chimaeric but, if you are lucky (and if
you've gotten this far, you obviously are),
its germ cells will carry the deleted gene.
A few rounds of careful breeding can then
produce progeny in which both copies of the
gene are inactivated.
L-chain:
Immunoglobulin light chain.
Leader sequence: Two
usages;
1] N-terminal pre
sequence of scretory proteins such as
peptide hormones and membrane proteins.
2] The untranslated
sequence at the 5'-ends of mRNA molecules.
Leucine zipper: A
motif found in certain proteins in which Leu
residues are evenly spaced through an
a-helical region, such that they would end
up on the same face of the helix. Dimers can
form between two such proteins. The Leu
zipper is important in the function of
transcription factors such as Fos and Jun
and related proteins.
Library: A library
might be either a genomic library, or a cDNA
library. In either case, the library is just
a tube carrying a mixture of thousands of
different clones - bacteria or l phages.
Each clone carries an "insert" -
the cloned DNA.
A cDNA library is usually
just a mixture of bacteria, where each
bacteria carries a different plasmid.
Inserted into the plasmids (one per plasmid)
are thousands of different pieces of cDNA
(each typ. 500-5000 bp) copied from some
source of mRNA, for example, total liver
mRNA. The basic idea is that if you have a
large enough number of different
liver-derived cDNAs carried in those
bacteria, there is a 99% probability that a
cDNA copy of any given liver mRNA exists
somewhere in the tube. The real trick is to
find the one you want out of that mess - a
process called screening (see
"Screening").
A genomic library is
similar in concept to a cDNA library, but
differs in three major ways - 1) the library
carries pieces of genomic DNA (and so
contains introns and flanking regions, as
well as coding and untranslated); 2) you
need bacteriophage l or cosmids, rather than
plasmids, because... 3) the inserts are
usually 5-15 kb long (in a l library) or
20-40 kb (in a cosmid library). Therefore, a
genomic library is most commonly a tube
containing a mixture of l phages. Enough
different phages must be present in the
library so that any given piece of DNA from
the source genome has a 99% probability of
being present.
Ligase: An enzyme, T4
DNA ligase, which can link pieces of DNA
together. The pieces must have compatible
ends (both of them blunt, or else mutually
compatible sticky ends), and the ligation
reaction requires ATP.
Ligation: The process
of splicing two pieces of DNA together. In
practice, a pool of DNA fragments are
treated with ligase (see "Ligase")
in the presence of ATP, and all possible
splicing products are produced, including
circularized forms and end-to-end ligation
of 2, 3 or more pieces. Usually, only some
of these products are useful, and the
investigator must have some way of selecting
the desirable ones.
Linear Epitope: An
epitope formed by a series of amino acids
which are adjacent to each other within the
primary structure of the protein. Such
epitopes can be successfully modelled by
synthetic peptides, but comprise only a
small proportion of all epitopes. The
minimal epitope size is about 5 amino acid
residues. Also called a sequential epitope.
Linkage: The measure
of proximity of two or more markers (e.g.,
genes, RFLP markers) on a chromosome
determined by recombination events. The
closer together the markers are, the lower
the probability that they will be separated
during DNA repair or replication processes
(binary fission in prokaryotes, mitosis or
meiosis in eukaryotes), and hence the
greater the probability that they will be
inherited together.
Linkage map: A map of
the relative positions of genetic loci on a
chromosome, determined on the basis of how
often the loci are inherited together.
Distance is measured in centimorgans (cM).
Linker: A small piece
of synthetic double-stranded DNA which
contains something useful, such as a
restriction site. A linker might be ligated
onto the end of another piece of DNA to
provide a desired restriction site.
Lipofectin: A
commercially marketed liposome suspension
which is mixed with DNA or RNA to facilitate
uptake of the nucleic acid by eukaryotic
cells (see Transfection).
Localize:
Determination of the original position
(locus) of a gene or other marker on a
chromosome.
Locus (pl. loci): The
position on a chromosome of a gene or other
chromosome marker; also, the DNA at that
position. The use of locus is sometimes
restricted to mean regions of DNA that are
expressed. See gene expression.
Long(q) and short(p)
arms: The regions either side of the
centromere, a compact part of a chromosome,
are known as arms. As the centromere is not
in the centre of the chromosome, one arm is
longer than the other.
M
M13: A bacteriophage
which infects certain strains of E. coli.
The salient feature of this phage is that it
packages only a single strand of DNA into
its capsid. If the investigator has inserted
some heterologous DNA into the M13 genome,
copious quantities of single-stranded DNA
can subsequently be isolated from the phage
capsids. M13 is often used to generate
templates for DNA sequencing.
Macrorestriction map:
Map depicting the order of and distance
between sites at which restriction enzymes
cleave chromosomes.
Maintenance methylase:
An enzymatic activity responsible for
maintaining the patterns of methylation on
each strand of a DNA molecule after
replication.
Mapping: See gene
mapping, linkage map, physical map.
Marker: Two typical
usages:
1] Molecular weight size
marker: a piece of DNA of known size, or
a mixture of pieces with known size, used on
electrophoresis gels to determine the size
of unknown DNA's by comparison.
2] Genetic marker: An
identifiable physical location on a
chromosome (e.g., restriction enzyme cutting
site, gene) whose inheritance can be
monitored. Markers can be expressed regions
of DNA (genes) or some segment of DNA with
no known coding function but whose pattern
of inheritance can be determined. See RFLP,
restriction fragment length polymorphism.
Mb: See megabase.
Megabase (Mb): Unit
of length for DNA fragments equal to 1
million nucleotides and roughly equal to 1
cM.
Melting: The
dissociation of a duplex nucleic acid
molecule into single strands, usually by
increasing temperature. See Denaturation.
Meiosis: The process
of two consecutive cell divisions in the
diploid progenitors of sex cells. Meiosis
results in four rather than two daughter
cells, each with a haploid set (1n) of
chromosomes.
Messenger RNA (mRNA):
Proteins are not synthesized directly from
genomic DNA. Instead, an RNA template (a
precursor mRNA) is constructed from the
sequence of the gene. This RNA is then
processed in various ways, including
splicing. Spliced single-stranded RNAs
destined to become templates for protein
synthesis are known as mRNAs, The term mRNA
is used only for a mature transcript with
polyA tail and with all introns removed,
rather than the primary transcript in the
nucleus. As such, an mRNA will have a 5'
untranslated region, a coding region, a 3'
untranslated region and (almost always) a
poly(A) tail. Typically about 2% of the
total cellular RNA is mRNA. See genetic
code.
Molecular biology:
The biochemical study of the genetic basis
for phenotype.
Monocistronic: A form
of gene organization resulting in
transcription of an mRNA that contains
coding sequence for a single gene or gene
product.
mRNA: See messenger
RNA.
mRNA export: Refers
to the movement of spliced mRNA out of the
nucleus to the cytoplasm.
mRNA processing:
Refers to the processes of polyadenylation,
splicing, and addition of a 5' cap
structure.
Metaphase: A stage in
mitosis or meiosis during which the
chromosomes are aligned along the equatorial
plane of the cell.
Missense Mutation: A
nucleotide mutation which results in a
change in the amino acid sequence of the
encoded protein (contrast with Silent
Mutation).
Mitosis: The process
of nuclear division in cells that produces
daughter cells that are genetically
identical to each other and to the parent
cell.
Molecular Genetics:
Study of how genes function to control
cellular activities.
Monoclonal Antibody:
An antibody with very specific and often
unique binding specificity which is secreted
by a biologically cloned line of
plasmacytoma cells in the absence of other
related antibodies with different binding
specificities. Differs from polyclonal
antibodies, which are mixed populations of
antibody molecules such as may be present in
a serum specimen, within which many
different individual antibodies have
different binding specificities.
Motif: A recurring
pattern of short sequence of DNA, RNA, or
protein, that usually serves as a
recognition site or active site. The same
motif can be found in a variety of types of
organisms.
Multicistronic Message:
An mRNA transcript with more than one
cistron and thus encoding more than one
polypeptide. These generally do not occur in
eukaryotic organisms, due to differences in
the mechanism of translation initiation.
Multicopy Plasmids:
Present in bacteria at amounts greater than
one per chromosome. Vectors for cloning DNA
are usually multicopy; there are sometimes
advantages in using a single copy plasmid.
Multifactorial or
multigenic disorders: See polygenic
disorders.
Multi-gene family: A
group of genes that are related by sequence
homologies; usually are also related by
their functions or by the processes in which
they participate.
Multiple Cloning Site:
An artificially constructed region within a
vector molecule which contains a number of
closely spaced recognition sequences for
restriction endonucleases. This serves as a
convenient site into which foreign DNA may
be inserted.
Multiplexing: A
sequencing approach that uses several pooled
samples simultaneously, greatly increasing
sequencing speed.
Mutagen: An agent
capable of causing mutations. Common
examples are ultraviolet light, such as in
sunlight, and anthracene, a material formed
during the cooking of fatty meats on a
barbecue grill.
Mutation: A
permanent, heritable change of the genetic
material, either in a single gene or in the
numbers or structures of the chromosomes.
Mutations do not always have harmful
effects. Compare polymorphism.
Native Gel: An
electrophoresis gel run under conditions
which do not denature proteins (i.e., in the
absence of SDS, urea, 2-mercaptoethanol,
etc.).
Nested Pcr: A very
sensitive method for amplfication of DNA,
which takes part of the product of a single
PCR reaction (after 30-35 cycles), and
subjects it to a new round of PCR using a
different set of PCR primers which are
nested within the region flanked by the
original primer pair (see Polymerase Chain
Reaction).
Nick: In duplex DNA,
this refers to the absence of a
phosphodiester bond between two adjacent
nucleotides on one strand.
Nick translation: A
method for incorporating radioactive
isotopes (typically 32P) into a piece of
DNA. The DNA is randomly nicked by DNase I,
and then starting from those nicks DNA
polymerase I digests and then replaces a
stretch of DNA. Radiolabeled precursor
nucleotide triphosphates can thus be
incorporated.
Nitrogenous base: A
nitrogen- containing molecule having the
chemical properties of a base.
Non-coding strand:
Anti-sense strand. See "Sense
strand" for a discussion of sense
strand vs. anti-sense strand.
Nonconservative
Substitution: A mutation which results
in the substitution of one amino acid within
a polypeptide chain with an amino acid
belonging to a different polarity/charge
group (see Amino Acids, Conservative
Mutation)
Northern blot: A
technique for analyzing mixtures of RNA by
transfer of size-separated RNA fragments to
a synthetic membrane, whereby the presence
and rough size of one particular type of RNA
(usually an mRNA) can be ascertained. See
"Blotting" for more information.
After Dr. E. M. Southern invented the
Southern blot, it was adapted to RNA and
named the "Northern" blot.
Nonsence Codon: See
Stop Codon.
Nonsense Mutation: A
change in the sequence of a nucleic acid
that causes a nonsense (stop or termination)
codon to replace a codon representing an
amino acid.
Nontranslated RNA (NTR):
The segments located at the 5' and 3' ends
of a mRNA molecule which do not encode any
part of the polyprotein; may contain
important translational control elements.
nt: Abbreviation for
nucleotide; i.e. the monomeric unit from
which DNA or RNA are built. One can express
the size of a nucleic acid strand in terms
of the number of nucleotides in its chain;
hence 'nt' can be a measure of chain length.
N Terminus: See Amino
Terminus.
Nuclear run-on: A
method used to estimate the relative rate of
transcription of a given gene, as opposed to
the steady-state level of the mRNA
transcript (which is influenced not just by
transcription rates, but by the stability of
the RNA). This technique is based on the
assumption that a highly-transcribed gene
should have more molecules of RNA polymerase
bound to it than will the same gene in a
less-active state. If properly prepared,
isolated nuclei will continue to transcribe
genes and incorporate 32P into RNA, but only
in those transcripts that were in progress
at the time the nuclei were isolated. Once
the polymerase molecules complete the
transcript they have in progress, they
should not be able to re-initiate
transcription. If that is true, then the
amount of radiolabel incorporated into a
specific type of mRNA is theoretically
proportional to the number of RNA polymerase
complexes present on that gene at the time
of isolation. A very difficult technique,
rarely applied appropriately from what I
understand.
Nuclease: An enzyme
which degrades nucleic acids. A nuclease can
be DNA-specific (a DNase), RNA-specific
(RNase) or non-specific. It may act only on
single stranded nucleic acids, or only on
double-stranded nucleic acids, or it may be
non-specific with respect to strandedness. A
nuclease may degrade only from an end (an
exonuclease), or may be able to start in the
middle of a strand (an endonuclease). To
further complicate matters, many enzymes
have multiple functions; for example, Bal31
has a 3'-exonuclease activity on
double-stranded DNA, and an endonuclease
activity specific for single-stranded DNA or
RNA.
Nuclease protection
assay: See "RNase protection
assay".
Nucleic acid: A large
molecule composed of nucleotide subunits.
Nucleoside: A term
refering to the combination of adenine,
cytosine, guanine, or thymine with a ribose
or 2-deoxyribose sugar moiety. A nucleoside
is not phosphorylated.
Nucleotide: A
building block of DNA or RNA consisting of a
nitrogenous base (adenine, guanine, thymine,
or cytosine in DNA; adenine, guanine,
uracil, or cytosine in RNA), a phosphate
molecule, and a sugar molecule (deoxyribose
in DNA and ribose in RNA). Thousands of
nucleotides are linked to form a DNA or RNA
molecule. See DNA, base pair, RNA.
Nucleus: The cellular
organelle in eukaryotes that contains the
genetic material.
Oligodeoxyribonucleotide:
A short, single-stranded DNA molecule,
generally l5-50 nucleotides in length, which
may be used as a primer or a hybridization
probe. Oligodeoxyribonucleotides are
synthesized chemically under automated
conditions.
Oligonucleotide: See
Oligodeoxyribonucleotide.
Oncogene: A gene in a
tumor virus or in cancerous cells which,
when transferred into other cells, can cause
transformation (note that only certain cells
are susceptible to transformation by any one
oncogene). Functional oncogenes are not
present in normal cells. A normal cell has
many "proto-oncogenes" which serve
normal functions, and which under the right
circumstances can be activated to become
oncogenes. The prefix "v-"
indicates that a gene is derived from a
virus, and is generally an oncogene (like v-src
, v-ras, v-myb , etc). See also
"Transformation (with respect to
cultured cells)".
Oncogenic transformation:
A change in the behavioral phenotype of a
cell to one lacking in normal proliferative
control, and often involving invasive
characteristics.
Open reading frame:
That segment of a nucleic acid sequence that
lies between two stop codons, when
translated in a given reading frame. The
presence of an open reading frame is
necessary to encode a polypeptide sequence
(or an exon thereof), but the presence of an
open reading frame is not proof that a
polypeptide sequence is encoded in that
sequence. See "Reading frame" for
a simple example.
Operon: Genes, which
are grouped together for coordinate
regulation by the same regulator.
Origin of replication:
A site at which DNA replication is
initiated. There is only one in bacterial
chromosomes, but numerous origins in
eukaryotic chromosomal DNA. (Abbr.
"ori")
Origin recognition
complex: An organization of protein
factors assembled at an origin of
replication for the purpose of initiating
replication.
Overhang: A terminus
of a duplex DNA molecule which has one or
more unpaired nucleotides in one of the two
strands (hence either a 3' or 5' overhang).
Cleavage of DNA with many restriction
endonucleases leaves such overhangs (see
Sticky End).
Overlapping clones:
See genomic library.
Package: In
recombinant DNA procedures, refers to the
step of incorporation of cosmid or other
lambda vector DNA with an insert into a
phage head for transduction of DNA into
host.
Palindromic Sequence:
A nucleotide sequence which is the same when
read in either direction, usually consisting
of adjacent inverted repeats. Restriction
endonuclease recognition sites are
palindromes:
5'-->3'
GAATTC EcoRI recognition
site
CTTAAG
3'<--5'
Paratope: That region
within the antigen-binding site of an
immunoglobulin molecule responsible for
recognition of epitope structure.
pBR322: A common
plasmid. Along with the obligatory origin of
replication, this plasmid has genes which
make the E. coli host resistant to
ampicillin and tetracycline. It also has
several restriction sites (BamHI, PstI,
EcoRI, HindIII etc.) into which DNA
fragments could be spliced in order to clone
them.
PCR: See
polymerase_chain reaction.
Penetrance: Refers to
the proportion of individuals heterozygous
for a given dominant allele that express the
phenotype of that dominant allele.
Peptide: A molecule
formed by peptide bonds covalently linking
two or more amino acids. Short peptides
(generally less than 60 amino acid residues,
and usually only half that length) can be
chemically synthesized by one of several
different methods; larger peptides (more
correctly, polypeptides) are usually
expressed from recombinant DNA.
Peptide Bond: A
covalent bond between two amino acids, in
which the carboxyl group of one amino acid
(X1--COOH) and the amino group of an
adjacent amino acid (NH2--X2) react to form
X1-CO-NH-X2 plus H2O.
Peptide-binding groove:
That region of a Class I or II MHC molecule
that is responsible for binding processed
antigen peptides for presentation.
Peptidyl transferase:
An enzymatic activity of the ribosome
responsible for formation of the peptide
bond between the nascent polypeptide chain
and the amino acid carried by the charged
tRNA in the A site of the ribosome. In so
doing, the ribosome is moved along the mRNA
by one codon.
Phage: A virus for
which the natural host is a bacterial cell.
Used in the laboratory as a cloning vector.
(see bacteriophage)
Phagemid: A type of
plasmid which carries within its sequence a
bacteriophage replication origin (ori). When
the host bacterium is infected with
"helper" phage, the phagemid is
replicated along with the phage DNA and
packaged into phage capsids.
Phase variation:
Alternation in the type of flagellum
produced by a bacterium.
Phenotype: The
observable properties and physical
characteristics of of a cell or an organism
that is the result of its unique genotype.
Phosphodiester Bond:
The covalent bond between the 3' hydroxyl in
the sugar ring of one nucleotide and the 5'
phosphate group of the sugar ring of the
adjacent nucleotide residue within a nucleic
acid:
5'-Ribose- 3' - O - P(O)2
- O - 5' -Ribose - 3' - etc.
Phosphorylation: The
addition of a phosphate monoester to a
macromolecule, catalyzed by a specific
kinase enzyme. With respect to proteins,
certain amino acid side chains (serine,
threonine, tyrosine) are subject to
phosphorylation catalyzed by protein
kinases; altering the phosphorylation status
of a protein may have dramatic effects on
its biologic properties, and is a common
cellular control mechanism. With respect to
DNA, 5' ends must be phosphorylated for
ligation.
Physical map: A map
of the locations of identifiable landmarks
on DNA (e.g., restriction enzyme cutting
sites, genes), regardless of inheritance.
Distance is measured in base pairs. For the
human genome, the lowest- resolution
physical map is the banding patterns on the
24 different chromosomes; the highest-
resolution map would be the complete
nucleotide sequence of the chromosomes.
Plasmid: A class of
circular, autonomously replicating,
extrachromosomal DNA elements found in many
bacteria. Contain origins of replication to
ensure their maintenance. Some plasmids are
capable of integrating into the host genome.
A number of artificially constructed
plasmids are used as cloning vectors or to
alter the characteristics of the bacteria.
Common plasmids are pBR322, pGEM, pUC18.
Point Mutation: A
single nucleotide substitution within a
gene; there may be several point mutations
within a single gene. Point mutations do not
lead to a shift in reading frames, thus at
most cause only a single amino acid
substitution (see Frameshift Mutation).
Polyacrylamide Gel
(PAGE): Used to separate proteins and
smaller DNA fragments and oligonucleotides
by electrophoresis. When run under
conditions which denature proteins (i.e., in
the presence of 2-mercaptoethanol, SDS, and
possibly urea), molecules are separated
primarily on the basis of size.
PolyA tail: After an
mRNA is transcribed from a gene, the cell
adds a stretch of A residues (typically
50-200) to its 3' end. It is thought that
the presence of this "polyA tail"
increases the stability of the mRNA
(possibly by protecting it from nucleases).
Note that not all mRNAs have a polyA tail;
the histone mRNAs in particular do not.
Polycistronic: Refers
to a form of gene organization resulting in
transcription of an mRNA that contains the
coding sequences for multiple gene products,
each of which is independently translated
from the mRNA.
Polyclonal Antibody:
See Monoclonal Antibody.
Polygenic disorders:
Genetic disorders resulting from the
combined action of alleles of more than one
gene (e.g., heart disease, diabetes, and
some cancers). Although such disorders are
inherited, they depend on the simultaneous
presence of several alleles; thus the
hereditary patterns are usually more complex
than those of single- gene disorders.
Compare single- gene disorders.
Polymerase: An enzyme
which links individual nucleotides together
into a long strand, using another strand as
a template. There are two general types of
polymerase - DNA polymerases (which
synthesize DNA) and RNA polymerase (which
makes RNA). Within these two classes, there
are numerous sub-types of polymerase,
depending on what type of nucleic acid can
function as template and what type of
nucleic acid is formed. A DNA-dependant DNA
polymerase will copy one DNA strand starting
from a primer, and the product will be the
complementary DNA strand. A DNA-dependant
RNA polymerase will use DNA as a template to
synthesize an RNA strand.
Polymerase chain
reaction: A technique for replicating a
specific piece of DNA in vitro, even
in the presence of excess non-specific DNA.
Primers are added (which initiate the
copying of each strand) along with
nucleotides and heat stable Taq polymerase.
By cycling the temperature, the target DNA
is repetitively denatured and copied.
Because the newly synthesized DNA strands
can subsequently serve as additional
templates for the same primer sequences,
successive rounds of primer annealing,
strand elongation, and dissociation produce
rapid and highly specific amplification of
the desired sequence. PCR also can be used
to detect the existence of the defined
sequence in a DNA sample. A single copy of
the target DNA, even if mixed in with other
undesirable DNA, can be amplified to obtain
billions of replicates. PCR can be used to
amplify RNA sequences if they are first
converted to DNA via reverse transcriptase.
This two-phase procedure is known as
'RT-PCR'.
Polymerase Chain Reaction
(PCR) is the basis for a number of extremely
important methods in molecular biology. It
can be used to detect and measure
vanishingly small amounts of DNA and to
create customized pieces of DNA. It has been
applied to clinical diagnosis and therapy,
to forensics and to vast numbers of research
applications. It would be difficult to
overstate the importance of PCR to science.
Polymorphism:
Difference in DNA sequence among
individuals. To be called a polymorphism, a
variant should be present in a significant
number of people in the population. Genetic
variations occurring in more than 1% of a
population would be considered useful
polymorphisms for genetic linkage analysis.
Compare mutation.
Polynucleotide Kinase:
Enzyme which catalyzes the transfer of the
terminal phosphate of ATP to 5' hydroxyl
termini of polynucleotides, either DNAor
RNA. Usually derived from T4 bacteriophage.
Polypeptide: See
Peptide.
Polyprotein: A giant
polypeptide that contains multiple
individual protein sequences embedded within
it and which must be proteolytically cleaved
to yield the individual proteins.
Polysome: Complex of
mRNA and several ribosomes.
Post-transcriptional
regulation: Any process occurring after
transcription which affects the amount of
protein a gene produces. Includes RNA
processing efficiency, RNA stability,
translation efficiency, protein stability.
For example, the rapid degradation of an
mRNA will reduce the amount of protein
arising from it. Increasing the rate at
which an mRNA is translated will increase
the amount of protein product.
Post-translational
processing: The reactions which alter a
protein's covalent structure, such as
phosphorylation, glycosylation or
proteolytic cleavage.
Post-translational
regulation: Any process which affects
the amount of protein produced from a gene,
and which occurs AFTER translation in the
grand scheme of genetic expression.
Actually, this is often just a buzz-word for
regulation of the stability of the protein.
The more stable a protein is, the more it
will accumulate.
Primase: An
DNA-dependent RNA polymerase function that
synthesizes a short RNA primer used during
DNA replication by DNA polymerase. Primase
does not require a primer, only a template.
PRE: Progesterone
Response Element: A binding site in a
promoter to which the activated progesterone
receptor can bind. The progesterone receptor
is essentially a transcription factor which
is activated only in the presence of
progesterone . The activated receptor will
bind to a PRE, and transcription of the
adjacent gene will be altered. See also
"Response element".
pre-mRNA: An RNA
molecule which is transcribed from
chromosomal DNA in the nucleus of eukaryotic
cells, and subsequently processed through
splicing reactions to generate the mRNA
which directs protein synthesis in the
cytoplasm.
Primary Structure: Refers
to the sequence of amino acid residues or
nucleotides within protein or nucleic acid
molecules, respectively (also see Secondary
and Tertiary Structure).
Primary transcript:
When a gene is transcribed in the nucleus,
the initial product is the primary
transcript, an RNA containing copies of all
exons and introns. This primary transcript
is then processed by the cell to remove the
introns, to cleave off unwanted 3' sequence,
and to polyadenylate the 5' end. The mature
message thus formed is then exported to the
cytoplasm for translation.
Primer: A small
oligonucleotide (anywhere from 6 to 50 nt
long) used to prime DNA synthesis. The DNA
polymerases are only able to extend a
pre-existing strand along a template; they
are not able to take a naked single strand
and produce a complementary copy of it
de-novo. A primer which sticks to the
template is therefore used to initiate the
replication. Primers are necessary for DNA
sequencing and PCR.
Primer extension:
This is a method used to figure out how far
upstream from a fixed site the start of an
mRNA is. For example, perhaps you have
isolated a cDNA clone, but you don't think
that the clone has all of the 5'
untranslated region. To find out how much is
missing, you would first sequence the part
you have, and figure out which strand is
coding strand (usually the coding strand
will have a large open reading frame). Next,
you ask the DNA Synthesis Facility to make
an oligonucleotide complementary to the
5'-most region of the coding strand (and
thus complementary to the mRNA). This
"primer" is hybridized to mRNA
(say, a mixture of mRNA containing the one
in which you are interested), and reverse
transcriptase is added to copy the mRNA from
the primer out to the 5' end. The size of
the resulting DNA fragment shows how far
away from the 5' end your primer is.
Prion: An infectious
agent thought to be composed solely of
protein. Term is derived from "Protein
infectious agent".
Prokaryote: A
single-celled organism with a simple
internal structure and no nucleus. Bacteria
and archaebacteria are prokaryotes.
Promoter site: Region
of a DNA molecule 5' to a coding sequence
that is responsible for assembly of an RNA
polymerase complex and the initiation of
transcription.
Proof-reading:
Mechanism for correction of errors made
during synthesis of nucleic acids or
polypeptides by scrutiny of the products
after the nucleotides or amino acids have
already been incorporated.
Proteasome: A
specialized organelle within the cytoplasm
of a cell that is responsible for
degradation of cytoplasmically situated
proteins. The proteasome plays a key role in
normal protein turnover and in peptide
presentation by MHC Class I antigen.
Protein: The
complete, assembled form of a holoprotein,
containing all necessary subunits (eg.
hemoglobin is comprised of two a and two b
globin subunits, as well as a heme
prosthetic group) (see also polypeptide).
Proteome: The
complete set of proteins encoded by the
genome.
Probe: Single-
stranded DNA or RNA fragment of specific
base sequence, labeled either radioactively
(often incorporating 32P or 35S) or
immunologically, that are used to detect the
complementary base sequence by
hybridization. For example, if you want to
quantitate the levels of alpha subunit mRNA
in a preparation of pituitary RNA, you might
make a radiolabeled RNA in-vitro which is
complementary to the mRNA, and then use it
to probe a Northern blot of the pit RNA. A
probe can be radiolabeled, or tagged with
another functional group such as biotin. A
probe can be cloned DNA, or might be a
synthetic DNA strand. As an example of the
latter, perhaps you have isolated a protein
for which you wish to obtain a cDNA or
genomic clone. You might (pay to)
microsequence a portion of the protein,
deduce the nucleic acid sequence, (pay to)
synthesize an oligonucleotide carrying that
sequence, radiolabel it and use it as a
probe to screen a cDNA library or genomic
library. A better way is to call up someone
who already has the clone.
Prokaryote: Cell or
organism lacking a membrane- bound,
structurally discrete nucleus and other
subcellular compartments. Bacteria are
prokaryotes. Compare eukaryote. See
chromosomes.
Processing: The
reactions occurring in the nucleus which
convert the primary RNA transcript to a
mature mRNA. Processing reactions include
capping, splicing and polyadenylation. The
term can also refer to the processing of the
protein product, including proteolytic
cleavages, glycosylation, etc.
Promoter: The first
few hundred nucleotides of DNA
"upstream" (on the 5' side) of a
gene, which control the transcription of
that gene. The promoter is part of the 5'
flanking DNA, i.e. it is not transcribed
into RNA, but without the promoter, the gene
is not functional. Note that the definition
is a bit hazy as far as the size of the
region encompassed, but the
"promoter" of a gene starts with
the nucleotide immediately upstream from the
cap site, and includes binding sites for one
or more transcription factors which can not
work if moved farther away from the gene.
Protein: A large
molecule composed of one or more chains of
amino acids in a specific order; the order
is determined by the base sequence of
nucleotides in the gene coding for the
protein. Proteins are required for the
structure, function, and regulation of the
bodys cells, tissues, and organs, and each
protein has unique functions. Examples are
hormones, enzymes, and antibodies.
Proto-oncogene: A
gene present in a normal cell which carries
out a normal cellular function, but which
can become an oncogene under certain
circumstances. The prefix "c-"
indicates a cellular gene, and is generally
used for proto-oncogenes (examples: c-myb
, c-myc , c-fos , c-jun , etc).
Pseudogene: A region
of DNA that shows extensive similarity to a
known gene, but which cannot itself
function, either because it has lost the
signal required for transcription (the
promoter sequence) or because it carries
mutations that prevent it from being
translated into protein. It is generally
assumed that pseudogenes are copies of mRNA.
Pseudogenes are known in the globin system
but also in many other multigene families.
Pseudoknot: A feature
of RNA tertiary structure; best visualized
as two overlapping stem-loops in which the
loop of the first stem-loop participates as
half of the stem in the second stem-loop.
Pseudorevertant: A
mutant virus or organism which has recovered
a wildtype phenotype due to a second-site
mutation (potentially located in a different
region of the genome, or involving a
different polypeptide) which has eliminated
the effect of the initial mutation.
Pulsed field gel
electrophoresis (PFGE): A gel technique
which allows size-separation of very large
fragments of DNA, in the range of hundreds
of kb to thousands of kb. As in other gel
electrophoresis techniques, populations of
molecules migrate through the gel at a speed
related to their size, producing discrete
bands. In normal electrophoresis, DNA
fragments greater than a certain size limit
all migrate at the same rate through the
gel. In PFGE, the electrophoretic voltage is
applied alternately along two perpendicular
axes, which forces even the larger DNA
fragments to separate by size.
Purine: A nitrogen-
containing, single- ring, basic compound
that occurs in nucleic acids. The purines in
DNA and RNA are adenine and guanine.
Pyrimidine: A
nitrogen- containing, double- ring, basic
compound that occurs in nucleic acids. The
pyrimidines in DNA are cytosine and thymine;
in RNA, cytosine and uracil.
R
Random primed synthesis:
If you have a DNA clone and you want to
produce radioactive copies of it, one way is
to denature it (separate the strands), then
hybridize to that template a mixture of all
possible 6-mer oligonucleotides. Those
oligos will act as primers for the synthesis
of labeled strands by DNA polymerase (in the
presence of radiolabeled precursors).
Rare- cutter enzyme:
See restriction enzyme cutting site.
Reading frame: When
mRNA is translated by the cell, the
nucleotides are read three at a time. By
starting at different positions, the
groupings of three that are produced can be
entirely different. The following example
shows a DNA sequence and the three reading
frames in which it could be read. Not only
is an entirely different amino acid sequence
specified by the different reading frames,
but two of the three frames have stop
codons, and thus are not open reading frames
(asterisks indicate a stop codon).
A DNA open reading frame:
...ATG ACA TGT AAA GAT AGA CTA ACC TTT
TGG...
...Met Thr Cys Lys Asp
Arg Leu Thr Phe Trp...
Same bases, different
grouping: ...A TGA CAT GTA AAG ATA GAC TAA
CCT TTT GG...
... *** His Val Lys Ile
Asp *** Pro Phe Gly..
Same bases, another
grouping: ...AT GAC ATG TAA AGA TAG ACT AAC
CTT TTG G..
... Asp Met *** Arg ***
Thr Asn Leu Leu ...
If we shift the grouping
again, we will just get the first reading
frame again. The reading frame that is
actually used is determined by the first
methionine codon (the initiation codon).
Once that first AUG is recognized, the
pattern of triplet groupings follows
unambiguously.
Recessive: Allele
that determines phenotype only when
homozygous; does not affect phenotype when
heterozygous with a dominant allele.
Recombinant clones:
Clones containing recombinant DNA molecules.
See recombinant DNA technologies.
Recombinant DNA: The
combination of foreign DNA inserts with
vector DNA (e.g., plasmid, phage, cosmid,
etc.) to produce a clone within a host.
Recombinant DNA
technologies: Procedures used to join
together DNA segments in a cell- free system
(an environment outside a cell or organism).
Under appropriate conditions, a recombinant
DNA molecule can enter a cell and replicate
there, either autonomously or after it has
become integrated into a cellular
chromosome.
Recombination: The
process by which DNA is exchanged between
pairs of equivalent chromosomes (crossing
over) during egg and sperm formation.
Recombination has the effect of making the
chromosomes of the offspring distinct from
those of the parents.
Recombination-Repair:
A mode of filling a gap in one strand of
duplex DNA by retrieving a homologous single
strand from another duplex. Usually the
underlying mechanism behind homologous
recombination and gene conversion.
Recombinase:
Enzymatic activity responsible for
facilitating intra- or intermolecular
recombination of DNA molecules.
Regulatory regions or
sequences: A DNA base sequence that
controls gene expression.
Relaxed DNA: See
Supercoil.
Repetitive DNA: A
surprising portion of any genome consists
not of genes or structural elements, but of
frequently repeated simple sequences. These
may be short repeats just a few nt long,
like CACACA etc. They can also range up to a
few hundred nt long. Examples of the latter
include Alu repeats, LINEs, SINEs. The
function of these elements is often unknown.
In shorter repeats like di- and
tri-nucleotide repeats, the number of
repeating units can occasionally change
during evolution and descent. They are thus
useful markers for familial relationships
and have been used in paternity testing,
forensic science and in the identification
of human remains.
Replication: The act
of a cell making a copy of all or some part
its genomic DNA.
Replicon: A segment
of genomic DNA that contains an origin of
replication and is replicated under the
control of that origin.
Reporter Gene: The
use of a functional enzyme, such as
beta-galactosidase, luciferase, or
chloramphenicol acetyltransderase,
downstream of a gene, promoter, or
translational control element of interest,
to more easily identify successful
introduction of the gene into a host and to
measure transcription and/or translation.
Repression: A form of
gene regulation wherein the promoter is
prevented from assembling an RNA polymerase
complex, so that transcription does not
occur.
Resolution: Degree of
molecular detail on a physical map of DNA,
ranging from low to high.
Response element: By
definition, a "response element"
is a portion of a gene which must be present
in order for that gene to respond to some
hormone or other stimulus. Response elements
are binding sites for transcription factors.
Certain transcription factors are activated
by stimuli such as hormones or heat shock. A
gene may respond to the presence of that
hormone because the gene has in its promoter
region a binding site for hormone-activated
transcription factor. Example: the
glucocorticoid response element (GRE).
Residue: As applied
to proteins, what remains of an amino acid
after its incorporation into a peptide
chain, with subsequent loss of a water
molecule (see Peptide Bond).
Restriction: To
"restrict" DNA means to cut it
with a restriction enzyme. See
"Restriction Enzyme".
Restriction enzymes :
An endonuclease enzyme, isolated from
bacteria, that recognizes specific base-pair
sequences within DNA and causes
endonucleolytic cleavage of the DNA at a
site determined by the recognized DNA
sequences. The sites vary from frequent to
rare cutting, depending upon the length of
the restriction site.
For example, the
restriction enzyme BamHI locates and cuts
any occurrence of:
5'-GGATCC-3'
||||||
3'-CCTAGG-5'
Note that both strands
contain the sequence GGATCC, but in
antiparallel orientation. The recognition
site is thus said to be palindromic, which
is typical of restriction sites. Every copy
of a plasmid is identical in sequence, so if
BamHI cuts a particular circular plasmid at
three sites producing three
"restriction fragments", then a
million copies of that plasmid will produce
those same restriction fragments a million
times over.
Bacteria produce
restriction enzymes for protection against
invasion by foreign DNA such as phages. The
bacteria's own DNA is modified in such a way
as to prevent it from being clipped.
Bacteria contain over 600 such enzymes that
recognize and cut over 100 different DNA
sequences. See restriction enzyme cutting
site.
Restriction enzyme
cutting site: A specific nucleotide
sequence of DNA at which a particular
restriction enzyme cuts the DNA. Some sites
occur frequently in DNA (e.g., every several
hundred base pairs), others much less
frequently (rare-cutter; e.g., every 10,000
base pairs).
Restriction fragment:
The piece of DNA released after restriction
digestion of plasmids or genomic DNA. See
"Restriction enzyme". One can
digest a plasmid and isolate one particular
restriction fragment (actually a set of
identical fragments). The term also
describes the fragments detected on a
genomic blot which carry the gene of
interest.
Restriction map: A
"cartoon" depiction of the
locations within a stretch of known DNA
where restriction enzymes will cut.
The map usually indicates
the approximate length of the entire piece
(scale on the bottom), as well as the
position within the piece at which
designated enzymes will cut. This map
happens to be of a plasmid, and the two ends
are joined together with about 25 nt between
the EcoRI and HindIII sites.
Restriction site: See
Restriction enzyme.
Restriction fragment
length polymorphism (RFLP): Variation in
the distance between restriction enzyme
cleavage sites that exist within a
population producing unique DNA fingerprint
patterns.
Restriction fragment
length polymorphism (RFLP): Variation
between individuals in DNA fragment sizes
cut by specific restriction enzymes;
polymorphic sequences that result in RFLPs
are used as markers on both physical maps
and genetic linkage maps. RFLPs are usually
caused by mutation at a cutting site. See
marker.
Reticulocyte Lysate:
A lysate of rabbit reticulocytes, which has
been extensively digested with micrococcal
nuclease to destroy the reticulocyte mRNAs.
With the addition of an exogenous, usually
synthetic, mRNA, amino acids and a source of
energy (ATP), the translational machinery of
the reticulocyte (ribosomes, eukaryotic
translation factors, etc.) will permit in
vitro translation of the added mRNA with
production of a new polypeptide. This is
only one of several available in vitro
translation systems.
Retrovirus: Singel
stranded RNA virus which is replicated and
expressed via a double stranded DNA
intermediate. Retroviruses contain enzyme
reverse transcriptase (RNA dependent DNA
polymerase); this enzyme converts viral RNA
into DNA which can combine with DNA of host
cell and produce more viral particles.
Revertant: See Back
Mutation.
RFLP: Restriction
fragment length polymorphism; the acronym is
pronounced "riflip". Although two
individuals of the same species have almost
identical genomes, they will always differ
at a few nucleotides. Some of these
differences will produce new restriction
sites (or remove them), and thus the banding
pattern seen on a genomic Southern will thus
be affected. For any given probe (or gene),
it is often possible to test different
restriction enzymes until you find one which
gives a pattern difference between two
individuals - a RFLP. The less related the
individuals, the more divergent their DNA
sequences are and the more likely you are to
find a RFLP.
RFLP: See restriction
fragment length polymorphism.
Reverse transcriptase:
An enzyme found in the retroviruses which
catalyzes the RNA-dependent polymerization
of DNA (DNA copy from RNA template). RT is
used to make cDNA; one begins by isolating
polyadenylated mRNA, providing oligo-dT as a
primer, and adding nucleotide triphosphates
and RT to copy the RNA into cDNA.
Ribonuclease: see
"RNAse".
Ribonucleic acid (RNA):
A chemical found in the nucleus and
cytoplasm of cells; it plays an important
role in protein synthesis and other chemical
activities of the cell. The structure of RNA
is similar to that of DNA. There are several
classes of RNA molecules, including
messenger RNA, transfer RNA, ribosomal RNA,
and other small RNAs, each serving a
different purpose.
Ribonucleotides: See
nucleotide.
Riboprobe: A strand
of RNA synthesized in-vitro (usually
radiolabeled) and used as a probe for
hybridization reactions. An RNA probe can be
synthesized at very high specific activity,
is single stranded (and therefore will not
self anneal), and can be used for very
sensitive detection of DNA or RNA.
Ribosome: The large,
multi-subunit ribonucleoprotein cellular
complex responsible for translation of mRNA
into polypeptide sequences. Ribosomes are a
complex consisting of ribosomal RNAs (rRNA)
and several proteins.
Ribosomal Binding
Sequence (Shine-Dalgarno sequence): In
prokaryotic organisms, part or all of the
polypurine sequence AGGAGG located on mRNA
just upstream of an AUG initiation codon; it
is complementary to the sequence at the 3'
end of 16S rRNA; and involved in binding of
the ribosome to mRNA. The internal ribosomal
entry site found in some viruses may be an
analogous eukaryotic genetic element.
Ribosome binding site:
Sequences contained in an mRNA that organize
the assembly of a ribosome to initiate
translation of the mRNA into polypeptide.
Ribosomal RNA (rRNA):
A class of RNA found in the ribosomes of
cells.
Ribozyme: A
catalytically active RNA. A good example is
the hepatitis delta virus RNA which is
capable of self-cleavage and self-ligation
in the absence of protein enzymes.
RNA: See ribonucleic
acid.
RNase: Ribonuclease;
an enzyme which degrades RNA. It is
ubiquitous in living organisms and is
exceptionally stable. The prevention of
RNase activity is the primary problem in
handling RNA. There are many different
RNases, some of the more important include:
RNase A Cleaves ssRNA 3'
of pyrimidines
RNase T1 Cleaves ssRNA at
guanasine nucleotides
RNase V1 Cleaves dsRNA
(helical regions)
RNase H Degrades the RNA
part of RNA:DNA hybrids.
RNA polymerase:
Enzymatic activity responsible for
DNA-dependent synthesis of RNA. In
prokaryotes there is only one RNA
polymerase. In eukaryotes, there are three,
each of which transcribes a different group
of genes.
RNase protection assay:
This is a sensitive method to determine (1)
the amount of a specific mRNA present in a
complex mixture of mRNA and/or (2) the sizes
of exons which comprise the mRNA of
interest. A radioactive DNA or RNA probe (in
excess) is allowed to hybridize with a
sample of mRNA (for example, total mRNA
isolated from tissue), after which the
mixture is digested with single-strand
specific nuclease. Only the probe which is
hybridized to the specific mRNA will escape
the nuclease treatment, and can be detected
on a gel. The amount of radioactivity which
was protected from nuclease is proportional
to the amount of mRNA to which it
hybridized. If the probe included both
intron and exons, only the exons will be
protected from nuclease and their sizes can
be ascertained on the gel.
RNA Splicing: A
complex and incompletly understood series of
reactions occuring in the nucleus of
eukaryotic cells in which pre-mRNA
transcribed from chromosomal DNA is
processed such that noncoding regions of the
pre-mRNA (introns) are excised, and coding
regions (exons) are covalently linked to
produce an mRNA molecule ready for transport
to the cytoplasm. Because of splicing,
eukaryotic DNA representing a gene encoding
any given protein is usually much larger
than the mRNA from which the protein is
actually translated.
rRNA: "ribosomal
RNA"; any of several RNAs which become
part of the ribosome, and thus are involved
in translating mRNA and synthesizing
proteins. They are the most abundant RNA in
the cell (on a mass basis).
RT-PCR: See
'Polymerase Chain Reaction and reverse
transcriptase'.
S1 end mapping: A
technique to determine where the end of an
RNA transcript lies with respect to its
template DNA (the gene). Can't be described
in a short paragraph. See "RNAse
Protection assay" for a closely related
technique.
S1 nuclease: An
enzyme which digests only single-stranded
nucleic acids.
Screening: To screen
a library (see "Library") is to
select and isolate individual clones out of
the mixture of clones. For example, if you
needed a cDNA clone of the pituitary
glycoprotein hormone alpha subunit, you
would need to make (or buy) a pituitary cDNA
library, then screen that library in order
to detect and isolate those few bacteria
carrying alpha subunit cDNA.
There are two methods of
screening which are particularly worth
describing: screening by hybridization, and
screening by antibody.
Screening by
hybridization involves spreading the mixture
of bacteria out on a dozen or so agar plates
to grow several ten thousand isolated
colonies. Membranes are laid onto each
plate, and some of the bacteria from each
colony stick, producing replicas of each
colony in their original growth position).
The membranes are lifted and the adherent
bacteria are lysed, then hybridized to a
radioactive piece of alpha DNA (the source
of which is a story in itself - see
"Probe"). When X-ray film is laid
on the filter, only colonies carrying alpha
sequences will "light up". Their
position on the membranes show where they
grew on the original plates, so you now can
go back to the original plate (where the
remnants of the colonies are still alive),
pick the colony off the plate and grow it
up. You now have an unlimited source of
alpha cDNA.
Screening by antibody is
an option if the bacteria and plasmid are
designed to express proteins from the cDNA
inserts (see "Expression clones").
The principle is similar to hybridization,
in that you lift replica filters from
bacterial plates, but then you use the
antibody (perhaps generated after olde tyme
protein purification rituals) to show which
colony expresses the desired protein.
SDS: Sodium dodecyl
sulphate. It is an ionic detergent.
SDS-PAGE: Denaturing
protein gel electrophoresis (see
Polyacrylamide Gel Electrophoresis).
Secondary Structure:
(also see Primary and Tertiary Structure)
Local structure within a protein which is
conferred by the nature of the side chains
of adjacent amino acids (e.g., alpha helix,
beta sheet, random coil); local structure
within an RNA molecule which is conferred by
base pairing of nucleotides which are
relatively closely positioned within the
sequence (e.g., hairpins, stem-loop
structures).
Selection: The use of
particular conditions, such as the presence
of ampicillin, to allow survival only of
cells with a particular phenotype, such as
production of beta-lactamase.
Sense strand: A gene
has two strands: the sense strand and the
anti-sense strand. The Sense strand is, by
definition, the same 'sense' as the mRNA;
that is it can be translated exactly as the
mRNA sequence can. Given a sense strand with
the following sequence:
5' - ATG GGG CCA CGG CTG
TGA - 3'
Met Gly Pro Arg Leu stop
The anti-sense strand
will read as follows (note that the strand
has been reversed and complemented):
5' - TCA CAG CCG TGG CCC
CAT - 3'
The duplex DNA will pair
as follows:
5' - ATGGGGCCACGGCTGTGA -
3'
||||||||||||||||||
3' - TACCCCGGAGCCGACACT -
5'
Note however that when
the RNA is transcribed from this sequence,
the ANTI-SENSE strand is used as the
template for RNA polymerization. After all,
the RNA must base-pair with its template
strand (see Figure 3), so the process of
transcription produces the complement of the
anti-sense strand. This introduces some
confusion about terminology:
Some people use the term
'coding strand' and 'non-coding strand' to
refer to the sense and antisense strands,
respectively. Unfortunately, many people
interpret these terms in exactly the
opposite way. I consider the terms 'coding
strand' and 'non-coding strand' to be too
ambiguous. Some people use the exact
opposite definition for 'sense' and
'anti-sense' that I have given here. Be
aware of the possibility of a discrepancy.
Textbooks I have consulted generally agree
with the nomenclature given herein, albeit
some avoid defining these terms at all.
Sequence: As a noun,
the sequence of a DNA is a buzz word for the
structure of a DNA molecule, in terms of the
sequence of bases it contains. As a verb,
"to sequence" is to determine the
structure of a piece of DNA; i.e. the
sequence of nucleotides it contains.
Sequence homology:
Similarities in nucleic acid sequence and
organization, and in their encoded products,
that are sufficiently great as to imply
common ancestral origins.
Sequence Polymorphism:
See Polymorphism.
Sequence similarity:
Similarity in nucleic acid or polypeptide
sequences, particularly in shorter segments,
that may not be sufficient to imply common
ancestral origins.
Sequence tagged site
(STS): Short (200 to 500 base pairs) DNA
sequence that has a single occurrence in the
human genome and whose location and base
sequence are known. Detectable by polymerase
chain reaction, STSs are useful for
localizing and orienting the mapping and
sequence data reported from many different
laboratories and serve as landmarks on the
developing physical map of the human genome.
Expressed sequence tags (ESTs) are STSs
derived from cDNAs. (See EST)
Sequential Epitope:
See Linear Epitope.
Sequencing:
Determination of the order of nucleotides
(base sequences) in a DNA or RNA molecule or
the order of amino acids in a protein.
Sex chromosomes: The
X and Y chromosomes in human beings that
determine the sex of an individual. Females
have two X chromosomes in diploid cells;
males have an X and a Y chromosome. The sex
chromosomes comprise the 23rd chromosome
pair in a karyotype. Compare autosome.
Sex-linked: A trait
transmitted by one of the chromosomes
determining sex of an individual.
Short arm (p): One of
the two prominent segments of a chromosome;
the long or "q" arm is the other.
The arms of a given chromosome join at its
centromere.
Shotgun cloning: The
practice of randomly clipping a larger DNA
fragment into various smaller pieces,
cloning everything, and then studying the
resulting individual clones to figure out
what happened. For example, if one was
studying a 50 kb gene, it "may" be
a bit difficult to figure out the
restriction map. By randomly breaking it
into smaller fragments and mapping those, a
master restriction map could be deduced. See
also Shotgun sequencing.
Shotgun sequencing: A
way of determining the sequence of a large
DNA fragment which requires little brain
power but lots of late nights. The large
fragment is shotgun cloned (see above), and
then each of the resulting smaller clones
("subclones") is sequenced. By
finding out where the subclones overlap, the
sequence of the larger piece becomes
apparent. Note that some of the regions will
get sequenced several times just by chance.
Shuttle vector: A
type of cloning vector that contains
sequences enabling it to be propagated and
maintained in more than one type of host
(e.g. E. coli and mammalian cells).
For this purpose shuttle vectors carry
different origin of DNA replication which
are characteristic of the desired host
systems.
Sigma Factor: Subunit
of bacterial RNA polymerase which controls
the correct initiation of transcription.
These proteins increase the binding affinity
of RNA polymerase to a promoter. Different
sigma factors recognize different promoter
sequences.
Signal recognition
particle (SRP): A chaperonin complex
responsible for arresting polypeptide
synthesis and facilitating the docking of a
ribosome to the endoplasmic reticulum
membrane. Normally, on ribosomes translating
polypeptides destined for insertion into or
across the endoplasmic reticulum membrane
become associated with SRP.
Signal Peptidase: An
enzyme present within the lumen of the
endoplasmic reticulum which proteolytically
cleaves a secreted protein at the site of a
signal sequence.
Signal Sequence: A
hydrophobic amino acid sequence which
directs a growing peptide chain to be
secreted into the endoplasmic reticulum.
Silent Mutation: A
nucleotide substitution (never a single
deletion or insertion) which does not alter
the amino acid sequence of an encoded
protein due to the degeneracy of the genetic
code. Such mutations usually involve the
third base (wobble position) of codons.
Single- gene disorder:
Hereditary disorder caused by a mutant
allele of a single gene (e.g., Duchenne
muscular dystrophy, retinoblastoma, sickle
cell disease). Compare polygenic disorders.
Single nucleotide
polymorphism (SNP): A polymorphism
caused by the change of a single nucleotide.
Most genetic variation between individual
humans is believed to be due to SNPs.
Single-strand
conformational polymorphism (SSCP):
Relies on secondary and tertiary structural
differences between denatured and rapidly
cooled amplified DNA fragments that differ
slightly in their DNA sequence; different
SSCP alleles are resolved on non-denaturing
acrylamide gels, usually at low temperature;
ability to resolve alleles depends on
conditions of electrophoresis.
Site-Directed
Mutagenesis: The introduction of a
mutation, usually a point mutation or an
insertion, into a particular location in a
cloned DNA fragment. This mutated fragment
may be used to "knock out" a gene
in the organism of interest by homologous
recombination.
Site-Specific
Recombination: Occurs between two
specific but not necessarily homologous
sequences. Usually catalyzed by enzymes not
involved in general or homologous
recombination.
Slot blot: Similar to
a dot blot, but the analyte is put onto the
membrane using a slot-shaped template. The
template produces a consistently shaped
spot, thus decreasing errors and improving
the accuracy of the analysis. See Dot blot.
snRNA: Small nuclear
RNA; forms complexes with proteins to form
snRNPs; involved in RNA splicing,
polyadenylation reactions, other unknown
functions (probably).
snRNP:
"snerps", Small Nuclear
RiboNucleoProtein particles, which are
complexes between small nuclear RNAs and
7-10 proteins, and which are involved in RNA
splicing and polyadenylation reactions.
Solution hybridization:
A method closely related to RNase protection
(see "Rnase protection assay").
Solution hybridization is designed to
measure the levels of a specific mRNA
species in a complex population of RNA. An
excess of radioactive probe is allowed to
hybridize to the RNA, then single-strand
specific nuclease is used to destroy the
remaining unhybridized probe and RNA. The
"protected" probe is separated
from the degraded fragments, and the amount
of radioactivity in it is proportional to
the amount of mRNA in the sample which was
capable of hybridization. This can be a very
sensitive detection method.
Somatic: Refers to
non-germline cells. Somatic cells may become
terminally differentiated with alterations
in their overall genetic compliment, because
they are not responsible for passing along
the organism's genetic material to the
offspring.
Somatic cells: Any
cell in the body except gametes and their
precursors.
Somatic hypermutation:
A very high frequency of mutational events
that occur in specific loci, such as the
variable segments of expressed
immunoglobulin genes. Somatic hypermutation
in immunoglobulin genes occurs after all
rearrangements have occurred and provide
additional potential for variation in
immunoglobulin structure.
Southern blot: The
transfer (by absorption) of size-separated
(by electrophoresis) DNA fragments to a
synthetic membrane whereby the presence and
rough size of one particular fragment of DNA
can be ascertained by detection of specific
base sequences ascertained by radiolabeled
complementary probes. Initially described by
E.M. Southern.
Southwestern Blot:
The binding of protein to a nucleic acid on
a matrix similar to what is done for
western, northern, and southern blots. This
technique is used to identify DNA binding
proteins and the recognition sites for these
proteins.
SP6 RNA Polymerase: A
bacteriophage RNA polymerase which is
commonly used to transcribe plasmid DNA into
RNA. The plasmid must contain an SP6
promoter upstream of the relevant sequence.
Splice sequence: A
sequence within an mRNA molecule that is the
site at which splicing occurs.
Splicing: The process
that removes introns (non-protein-coding
portions) from transcribed RNAs. Exons
(protein-coding portions) can also be
removed. Depending on which exons are
removed, different proteins can be made from
the same initial RNA or gene. Different
proteins created in this way are 'splice
variants' or 'alternatively spliced'.
Splicosome: A
ribonucleoprotein structure responsible for
splicing of primary transcripts.
Stable transfection:
A form of transfection experiment designed
to produce permanent lines of cultured cells
with a new gene inserted into their genome.
Usually this is done by linking the desired
gene with a "selectable" gene,
i.e. a gene which confers resistance to a
toxin (like G418, aka Geneticin). Upon
putting the toxin into the culture medium,
only those cells which incorporate the
resistance gene will survive, and
essentially all of those will also have
incorporated the experimenter's gene.
Start codon: That
codon at which translation of an mRNA
molecule begins. This is always an AUG
(encoding methionine) in eukaryotes, and
nearly always in prokaryotes. In prokaryotes
N-formyl-methionine is used to initiate
polypeptide synthesis.
Sticky ends: After
digestion of a DNA with certain restriction
enzymes, the ends left have one strand
overhanging the other to form a short
(typically 4 nt) single-stranded segment.
This overhang will easily re-attach to other
ends like it, and are thus known as
"sticky ends". For example, the
enzyme BamHI recognizes the sequence GGATCC,
and clips after the first G in each strand:
The overhangs thus
produced can still hybridize
("anneal") with each other, even
if they came from different parent DNA
molecules, and the enzyme ligase will then
covalently link the strands. Sticky ends
therefore facilitate the ligation of diverse
segments of DNA, and allow the formation of
novel DNA constructs.
Stop codon: That
codon at which translation of an mRNA
molecule into a polypeptide is terminated.
In the Universal Code this may be: UGA, UAG,
or UAA.
Streptavidin: A
bacterial analog of egg white avidin.
Stringency: A term
used to describe the conditions of
hybridization. By varying the conditions
(especially salt concentration and
temperature) a given probe sequence may be
allowed to hybridize only with its exact
complement (high stringency), or with any
somewhat related sequences (relaxed or low
stringency). Increasing the temperature or
decreasing the salt concentration will tend
to increase the selectivity of a
hybridization reaction, and thus will raise
the stringency.
STS: See sequence
tagged site.
Sub-cloning: If you
have a cloned piece of DNA (say, inserted
into a plasmid) and you need unlimited
copies of only a part of it, you might
"sub-clone" it. This involves
starting with several million copies of the
original plasmid, cutting with restriction
enzymes, and purifying the desired fragment
out of the mixture. That fragment can then
be inserted into a new plasmid for
replication. It has now been subcloned.
Supercoil:
Double-stranded circular DNA which is
twisted about itself. Commonly observed with
plasmids and circular viral DNA genomes
(such as that of hepatitis B virus). A nick
in one strand of the plasmid may remove the
twist, resulting in a relaxed, circular DNA
molecule. A complete break in the DNA puts
the plasmid in a linear form. Supercoils,
relaxed circular DNA, and linear DNA all
have different migration properties in
agarose gels, even though they contain the
same number of base pairs.
T7 RNA Polymerase: A
bacteriophage RNA polymerase which is
commonly used to transcribe plasmid DNA into
RNA. The plasmid must contain a T7 promoter
upstream of the relevant sequence.
Tandem repeat sequences:
Multiple copies of the same base sequence on
a chromosome; used as a marker in physical
mapping.
Taq polymerase: A DNA
polymerase isolated from the bacterium Thermophilis
aquaticus and which is very stable to
high temperatures. It is used in PCR
procedures and high temperature sequencing.
TATA box: A sequence
found in the promoter (part of the 5'
flanking region) of many genes. Deletion of
this site (the binding site of transcription
factor TFIID) causes a marked reduction in
transcription, and gives rise to
heterogeneous transcription initiation
sites.
Technology transfer:
The process of converting scientific
findings from research laboratories into
useful products by the commercial sector.
Telomerase: A
ribonucleoprotein complex that maintains the
repeat sequence structures at the telomeric
ends of chromosomes.
Telomere: The natural
distal end of a chromosome. Contain some
form of simple repeating sequence, usually
with a single stranded distal end that may
form a hairpin. These specialized structures
are involved in the replication and
stability of linear DNA molecules. See DNA
replication.
Terminator: A
sequence downstream from the 3' end of an
open reading frame that serves to halt
transcription by the RNA polymerase. In
bacteria these are commonly sequences that
are palindromic and thus capable of forming
hairpins. Sometimes termination requires the
action of a protein, such as Rho factor in
E. coli.
Tertiary Structure:
(also see Primary and Secondary Structure)
Refers to higher ordered structures
conferred on proteins or nucleic acids by
interactions between amino acid residues or
nucleotides which are not closely positioned
within the sequence (primary structure) of
the molecule.
Tet resistance: See
"Antibiotic resistance".
Thymine (T): One of
the pyrimidine bases found in DNA one member
of the base pair A- T (adenine- thymine).
(2,4-dihydroxy-5-methylpyrimidine).
Tissue-specific
expression: Gene function which is
restricted to a particular tissue or cell
type. For example, the glycoprotein hormone
alpha subunit is produced only in certain
cell types of the anterior pituitary and
placenta, not in lungs or skin; thus
expression of the glycoprotein hormone
alpha-chain gene is said to be
tissue-specific. Tissue specific expression
is usually the result of an enhancer which
is activated only in the proper cell type.
Tm: The midpoint of
the temperature range over which DNA is
melted or denatured by heat; the temperature
at which a duplex nucleic acid molecule is
50% melted into single strands, it is
dependent upon the number and proportion of
G-C base pairs as well as the ionic
conditions. Often referred to as a measure
of the thermal stability of a nucleic acid
probe:target sequence hybrid.
Topoisomerase: An
enzymatic activity responsible for relieving
excessive supercoiling in DNA.
Trans-: Located on
two physically dis-contiguous DNA molecules.
Transcription: The
process of copying a gene into RNA
transcript. This is the first step in the
expression of any gene. The resulting RNA,
if it codes for a protein, will be spliced,
polyadenylated, transported to the
cytoplasm, and by the process of translation
will produce the desired protein molecule,
although not all transcripts lead to
proteins. Compare translation.
Transcription factor:
A protein which controls the transcription
of genes. These usually bind to DNA as part
of their function (but not necessarily). A
transcription factor may be general (i.e.
acting on many or all genes in all tissues),
or tissue-specific (i.e. present only in a
particular cell type, and activating the
genes restricted to that cell type). Its
activity may be constitutive, or may depend
on the presence of some stimulus; for
example, the glucocorticoid receptor is a
transcription factor which is active only
when glucocorticoids are present.
Transcription start site:
The point in a DNA sequence at which
transcription of a gene into RNA begins.
Transcriptome: The
complete set of RNAs transcribed from a
genome.
Transduction: The
incorporation of a cellular gene into a
viral genome, that can then be introduced
into other cells.
Transfection: A
method by which foreign DNA may be put into
a cultured mammalian cell. Such experiments
are usually performed using cloned DNA
containing coding sequences and control
regions (promoters, etc) in order to test
whether the DNA will be expressed. Since the
cloned DNA may have been extensively
modified (for example, protein binding sites
on the promoter may have been altered or
removed), this procedure is often used to
test whether a particular modification
affects the function of a gene.
Transfer RNA (tRNA):
A class of RNA having structures with
triplet nucleotide sequences that are
complementary to the triplet nucleotide
coding sequences of mRNA. The role of tRNAs
in protein synthesis is to bond with amino
acids and transfer them to the ribosomes,
where proteins are assembled according to
the genetic code carried by mRNA.
Transformation: A
process by which the genetic material
carried by an individual cell is altered by
incorporation of exogenous DNA into its
genome. The cancerous alteration of
mammalian cells
Transformation (with
respect to cultured cells): A change in
cell morphology and behavior which is
generally related to carcinogenesis.
Transformed cells tend to exhibit
characteristics known collectively as the
"transformed phenotype" (rounded
cell bodies, reduced attachment dependence,
increased growth rate, loss of contact
inhibition, etc). There are different
"degrees" of transformation, and
cells may exhibit only a subset of these
characteristics. Not well understood, the
process of transformation is the subject of
intense research.
Transformed phenotype:
Acquisition of a phenotype characteristic of
oncogenic transformation.
Transgenic: Creation
of an individual in which genetic
modification has occured in the germline
tissues and may be transmitted to subsequent
generations.
Transgenic mouse: A
mouse which carries experimentally
introduced DNA. The
procedure by which one
makes a transgenic mouse involves the
injection of DNA into a fertilized embryo at
the pro-nuclear stage. The DNA is generally
cloned, and may be experimentally altered.
It will become incorporated into the genome
of the embryo. That embryo is implanted into
a foster mother, who gives birth to an
animal carrying the new gene. Various
experiments are then carried out to test the
functionality of the inserted DNA.
Transient transfection:
When DNA is transfected into cultured cells,
it is able to stay in those cells for about
2-3 days, but then will be lost (unless
steps are taken to ensure that it is
retained - see Stable transfection). During
those 2-3 days, the DNA is functional, and
any functional genes it contains will be
expressed. Investigators take advantage of
this transient expression period to test
gene function.
Transition: A
nucleotide substitution in which one
pyrimidine is replaced by the other
pyrimidine, or one purine replaced by the
other purine (e.g., A is changed to G, or C
is changed to T) (contrast with
Transversion) .
Translation: The
process of converting the genetic code into
polypeptides, catalyzed by the ribosome and
a host of soluble factors. mRNA codons are
recognized by tRNA anti-codons. Each tRNA
codes for a single amino acid, resulting in
synthesis of polypeptide wherein the amino
acid sequence is dictated by and matches the
order of the codons in the mRNA. Sometimes,
however, people speak of
"translating" the DNA or RNA when
they are merely reading the nucleotide
sequence and predicting from it the sequence
of the encoded protein. This might be more
accurately termed "conceptual
translation". Compare transcription.
Translational frame
shifting: A mechanism used by certain
viruses and even higher organisms to change
the reading frame used during translation of
an mRNA molecule, in a controlled manner, so
that the polypeptide product is the result
of translation in more than one reading
frame.
Translocation: The
process by which a newly synthesized protein
is directed toward a specific cellular
compartment (i.e, the nucleus, the
endoplasmic reticulum).
Transposible elements:
Genetic elements characterized by their
abilities to insert into and withdraw from a
given location within the genome, resulting
in movement from site to site within the
genome over a period of time. Transposable
elements may cause epigenetic changes in
phenotype.
Transposition: The
movement of DNA from one location to another
location on the same molecule, or a
different molecule within a cell.
Transversion: A
nucleotide substitution in which a purine
replaces a pyrimidine, or vice versa (e.g.,
A is changed to T, or T is changed to G)
(see Transition)
tRNA (transfer RNA):
Special RNAs that are charged with amino
acids and which carry anticodons for
recognition of the codons in mRNA. tRNAs are
responsible for translation from the
language of nucleic acids into the language
of polypeptides. Each tRNA can be charged
with only one type of amino acid, although
multiple tRNAs exist for many of the amino
acids.
Tumor suppressor: A
gene that prevents tumor formation until
deleted or mutated. The best-known examples
of tumor suppressors are the proteins p53
and Rb.
Turnover: The balance
between synthesis and degradation of a
product.
Universal Genetic Code:
The "standard" codon usage that is
common to most organisms.
Untranslated RNA: See
Nontranslated RNA.
Upstream activator
sequence: A binding site for
transcription factors, generally part of a
promoter region. A UAS may be found upstream
of the TATA sequence (if there is one), and
its function is (like an enhancer) to
increase transcription. Unlike an enhancer,
it can not be positioned just anywhere or in
any orientation.
Upstream/Downstream:
In an RNA, anything towards the 5' end of a
reference point is "upstream" of
that point. This orientation reflects the
direction of both the synthesis of mRNA, and
its translation - from the 5' end to the 3'
end. In DNA, the situation is a bit more
complicated. In the vicinity of a gene (or
in a cDNA), the DNA has two strands, but one
strand is virtually a duplicate of the RNA,
so it's 5' and 3' ends determine upstream
and downstream, respectively. NOTE that in
genomic DNA, two adjacent genes may be on
different strands and thus oriented in
opposite directions. Upstream or downstream
is only used on conjunction with a given
gene.
Uracil: A nitrogenous
base normally found in RNA but not DNA;
uracil is capable of forming a base pair
with adenine.
Vector: A construct
used to propagate DNA in a host (bacteria,
yeast, or cultured cells). The vector
provides all sequences essential for
replicating the test DNA. Typical vectors
include plasmids, cosmids, phages and YACs.
Virion: A
replication-competent virus particle.
Virulence: Ability to
infect or cause disease.
Virus: A noncellular
biological entity that can reproduce only
within a host cell. Viruses consist of
nucleic acid covered by protein; some animal
viruses are also surrounded by membrane.
Inside the infected cell, the virus uses the
synthetic capability of the host to produce
progeny virus.
VLSI: Very large-
scale integration allowing over 100,000
transistors on a chip.
Western blot: A
technique for analyzing mixtures of proteins
to show the presence, size and abundance of
one particular type of protein. Similar to
Southern or Northern blotting (see
"Blotting"), except that (1) a
protein mixture is electrophoresed in an
acrylamide gel, and (2) the
"probe" is an antibody which
recognizes the protein of interest, followed
by a radioactive secondary probe (such as
125I-protein A).
Wildtype: The native
or predominant genetic constitution before
mutations, usually referring to the genetic
consitution normally existing in nature.
Wobble Position: The
third base position within a codon, which
can often (but not always) be altered to
another nucleotide without changing the
encoded amino acid (see Degeneracy).
Xeno-immunization:
Stimulation of immune response to antigens
from different species.
YAC: See yeast
artificial chromosome.
Yeast artificial
chromosome (YAC): This is a method for
cloning very large fragments of DNA. Genomic
DNA in fragments of 200-500 kb are linked to
sequences which allow them to propagate in
yeast as a mini-chromosome (including
telomeres, a centromere and an ARS - an
autonomous replication sequence). This
technique is used to clone large genes and
intergenic regions, and for chromosome
walking. Compare cloning vector, cosmid.
Z-DNA: Alternative
left-handed form of the double helix which
retains Watson-Crick type base pairing.
Zinc finger: A
protein structural motif common in DNA
binding proteins. Four Cys residues are
found for each "finger" and one
finger can bind a molecule of zinc. A
typical configuration is:
CysXxxXxxCys--(intervening 12 or so
aa's)--CysXxxXxxCys.