Molecular biology /məˈlɛkjʊlər/ is a branch of biochemistry which
concerns the molecular basis of biological activity between
biomolecules in the various systems of a cell, including the
interactions between DNA, RNA, and proteins and their biosynthesis, as
well as the regulation of these interactions. Writing in Nature in
William Astbury described molecular biology as:
"...not so much a technique as an approach, an approach from the
viewpoint of the so-called basic sciences with the leading idea of
searching below the large-scale manifestations of classical biology
for the corresponding molecular plan. It is concerned particularly
with the forms of biological molecules and [...] is predominantly
three-dimensional and structural—which does not mean, however, that
it is merely a refinement of morphology. It must at the same time
inquire into genesis and function."
1 Relationship to other biological sciences
2 Techniques of molecular biology
2.1 Molecular cloning
2.2 Polymerase chain reaction
2.3 Gel electrophoresis
2.4 Macromolecule blotting and probing
2.4.1 Southern blotting
2.4.2 Northern blotting
2.4.3 Western blotting
2.4.4 Eastern blotting
2.6 Allele-specific oligonucleotide
4 See also
6 Further reading
7 External links
Relationship to other biological sciences
Schematic relationship between biochemistry, genetics and molecular
Researchers in molecular biology use specific techniques native to
molecular biology but increasingly combine these with techniques and
ideas from genetics and biochemistry. There is not a defined line
between these disciplines. The figure to the right is a schematic that
depicts one possible view of the relationships between the fields:
Biochemistry is the study of the chemical substances and vital
processes occurring in live organisms. Biochemists focus heavily on
the role, function, and structure of biomolecules. The study of the
chemistry behind biological processes and the synthesis of
biologically active molecules are examples of biochemistry.
Genetics is the study of the effect of genetic differences on
organisms. This can often be inferred by the absence of a normal
component (e.g. one gene). The study of "mutants" – organisms which
lack one or more functional components with respect to the so-called
"wild type" or normal phenotype. Genetic interactions (epistasis) can
often confound simple interpretations of such "knockout" studies.
Molecular biology is the study of molecular underpinnings of the
processes of replication, transcription, translation, and cell
function. The central dogma of molecular biology where genetic
material is transcribed into
RNA and then translated into protein,
despite being oversimplified, still provides a good starting point for
understanding the field. The picture has been revised in light of
emerging novel roles for RNA.
Much of molecular biology is quantitative, and recently much work has
been done at its interface with computer science in bioinformatics and
computational biology. In the early 2000s, the study of gene structure
and function, molecular genetics, has been among the most prominent
sub-fields of molecular biology. Increasingly many other areas of
biology focus on molecules, either directly studying interactions in
their own right such as in cell biology and developmental biology, or
indirectly, where molecular techniques are used to infer historical
attributes of populations or species, as in fields in evolutionary
biology such as population genetics and phylogenetics. There is also a
long tradition of studying biomolecules "from the ground up" in
Techniques of molecular biology
For more extensive list on protein methods, see protein methods. For
more extensive list on nucleic acid methods, see nucleic acid methods.
Main article: Molecular cloning
One of the most basic techniques of molecular biology to study protein
function is molecular cloning. In this technique,
DNA coding for a
protein of interest is cloned using polymerase chain reaction (PCR),
and/or restriction enzymes into a plasmid (expression vector). A
vector has 3 distinctive features: an origin of replication, a
multiple cloning site (MCS), and a selective marker usually antibiotic
resistance. Located upstream of the multiple cloning site are the
promoter regions and the transcription start site which regulate the
expression of cloned gene. This plasmid can be inserted into either
bacterial or animal cells. Introducing
DNA into bacterial cells can be
done by transformation via uptake of naked DNA, conjugation via
cell-cell contact or by transduction via viral vector. Introducing DNA
into eukaryotic cells, such as animal cells, by physical or chemical
means is called transfection. Several different transfection
techniques are available, such as calcium phosphate transfection,
electroporation, microinjection and liposome transfection. The plasmid
may be integrated into the genome, resulting in a stable transfection,
or may remain independent of the genome, called transient
DNA coding for a protein of interest is now inside a cell, and the
protein can now be expressed. A variety of systems, such as inducible
promoters and specific cell-signaling factors, are available to help
express the protein of interest at high levels. Large quantities of a
protein can then be extracted from the bacterial or eukaryotic cell.
The protein can be tested for enzymatic activity under a variety of
situations, the protein may be crystallized so its tertiary structure
can be studied, or, in the pharmaceutical industry, the activity of
new drugs against the protein can be studied.
Polymerase chain reaction
Main article: Polymerase chain reaction
Polymerase chain reaction
Polymerase chain reaction (PCR) is an extremely versatile technique
for copying DNA. In brief,
PCR allows a specific
DNA sequence to be
copied or modified in predetermined ways. The reaction is extremely
powerful and under perfect conditions could amplify one
to become 1.07 billion molecules in less than two hours. The PCR
technique can be used to introduce restriction enzyme sites to ends of
DNA molecules, or to mutate particular bases of DNA, the latter is a
method referred to as site-directed mutagenesis.
PCR can also be used
to determine whether a particular
DNA fragment is found in a cDNA
PCR has many variations, like reverse transcription PCR
(RT-PCR) for amplification of RNA, and, more recently, quantitative
PCR which allow for quantitative measurement of
DNA or RNA
Main article: Gel electrophoresis
Two percent Agarose Gel in Borate Buffer cast in a Gel Tray (Front,
Gel electrophoresis is one of the principal tools of molecular
biology. The basic principle is that DNA, RNA, and proteins can all be
separated by means of an electric field and size. In agarose gel
RNA can be separated on the basis of size by
DNA through an electrically charged agarose gel. Proteins
can be separated on the basis of size by using an
SDS-PAGE gel, or on
the basis of size and their electric charge by using what is known as
a 2D gel electrophoresis.
Macromolecule blotting and probing
The terms northern, western and eastern blotting are derived from what
initially was a molecular biology joke that played on the term
Southern blotting, after the technique described by
Edwin Southern for
the hybridisation of blotted DNA. Patricia Thomas, developer of the
RNA blot which then became known as the northern blot, actually didn't
use the term.
Main article: Southern blot
Named after its inventor, biologist Edwin Southern, the Southern blot
is a method for probing for the presence of a specific
DNA samples before or after restriction enzyme
(restriction endonuclease) digestion are separated by gel
electrophoresis and then transferred to a membrane by blotting via
capillary action. The membrane is then exposed to a labeled
that has a complement base sequence to the sequence on the
interest. Southern blotting is less commonly used in laboratory
science due to the capacity of other techniques, such as PCR, to
DNA sequences from
DNA samples. These blots are still
used for some applications, however, such as measuring transgene copy
number in transgenic mice or in the engineering of gene knockout
embryonic stem cell lines.
Main article: Northern blot
Northern blot diagram
The northern blot is used to study the expression patterns of a
specific type of
RNA molecule as relative comparison among a set of
different samples of RNA. It is essentially a combination of
RNA gel electrophoresis, and a blot. In this process
separated based on size and is then transferred to a membrane that is
then probed with a labeled complement of a sequence of interest. The
results may be visualized through a variety of ways depending on the
label used; however, most result in the revelation of bands
representing the sizes of the
RNA detected in sample. The intensity of
these bands is related to the amount of the target
RNA in the samples
analyzed. The procedure is commonly used to study when and how much
gene expression is occurring by measuring how much of that
present in different samples. It is one of the most basic tools for
determining at what time, and under what conditions, certain genes are
expressed in living tissues.
Main article: Western blot
In western blotting, proteins are first separated by size, in a thin
gel sandwiched between two glass plates in a technique known as
SDS-PAGE. The proteins in the gel are then transferred to a
polyvinylidene fluoride (PVDF), nitrocellulose, nylon, or other
support membrane. This membrane can then be probed with solutions of
antibodies. Antibodies that specifically bind to the protein of
interest can then be visualized by a variety of techniques, including
colored products, chemiluminescence, or autoradiography. Often, the
antibodies are labeled with enzymes. When a chemiluminescent substrate
is exposed to the enzyme it allows detection. Using western blotting
techniques allows not only detection but also quantitative analysis.
Analogous methods to western blotting can be used to directly stain
specific proteins in live cells or tissue sections.
Main article: Eastern blot
The eastern blotting technique is used to detect post-translational
modification of proteins. Proteins blotted on to the PVDF or
nitrocellulose membrane are probed for modifications using specific
DNA microarray being printed
Hybridization of target to probe
DNA microarray is a collection of spots attached to a solid support
such as a microscope slide where each spot contains one or more
DNA oligonucleotide fragment. Arrays make it possible
to put down large quantities of very small (100 micrometre diameter)
spots on a single slide. Each spot has a
DNA fragment molecule that is
complementary to a single
DNA sequence. A variation of this technique
allows the gene expression of an organism at a particular stage in
development to be qualified (expression profiling). In this technique
RNA in a tissue is isolated and converted to labeled cDNA. This
DNA is then hybridized to the fragments on the array and
visualization of the hybridization can be done. Since multiple arrays
can be made with exactly the same position of fragments they are
particularly useful for comparing the gene expression of two different
tissues, such as a healthy and cancerous tissue. Also, one can measure
what genes are expressed and how that expression changes with time or
with other factors. There are many different ways to fabricate
microarrays; the most common are silicon chips, microscope slides with
spots of ~100 micrometre diameter, custom arrays, and arrays with
larger spots on porous membranes (macroarrays). There can be anywhere
from 100 spots to more than 10,000 on a given array. Arrays can also
be made with molecules other than DNA.
Main article: Allele-specific oligonucleotide
Allele-specific oligonucleotide (ASO) is a technique that allows
detection of single base mutations without the need for
PCR or gel
electrophoresis. Short (20-25 nucleotides in length), labeled probes
are exposed to the non-fragmented target DNA, hybridization occurs
with high specificity due to the short length of the probes and even a
single base change will hinder hybridization. The target
DNA is then
washed and the labeled probes that didn't hybridize are removed. The
DNA is then analyzed for the presence of the probe via
radioactivity or fluorescence. In this experiment, as in most
molecular biology techniques, a control must be used to ensure
In molecular biology, procedures and technologies are continually
being developed and older technologies abandoned. For example, before
the advent of
DNA gel electrophoresis (agarose or polyacrylamide), the
DNA molecules was typically determined by rate sedimentation
in sucrose gradients, a slow and labor-intensive technique requiring
expensive instrumentation; prior to sucrose gradients, viscometry was
used. Aside from their historical interest, it is often worth knowing
about older technology, as it is occasionally useful to solve another
new problem for which the newer technique is inappropriate.[citation
Main article: History of molecular biology
While molecular biology was established in the 1930s, the term was
Warren Weaver in 1938. Weaver was the director of Natural
Sciences for the
Rockefeller Foundation at the time and believed that
biology was about to undergo a period of significant change given
recent advances in fields such as X-ray crystallography.
Clinical research and medical therapies arising from molecular biology
are partly covered under gene therapy. The use of molecular biology or
molecular cell biology approaches in medicine is now called molecular
Molecular biology also plays important role in understanding
formations, actions, and regulations of various parts of cells which
can be used to efficiently target new drugs, diagnosis disease, and
understand the physiology of the cell.
Central dogma of molecular biology
Molecular biology institutes
Protein interaction prediction
Protein structure prediction
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Biochemistry and Molecular
Biology at Curlie (based on DMOZ)
History of molecular biology
DNA replication (DNA)
Dry lab / Wet lab
Model organisms (such as
Pigment & Radioactivity
High-throughput technique ("-omics")
Molecular and cellular biology
DNA → RNA → Protein
RNA (pre-mRNA / hnRNA)
Histone acetylation and deacetylation
Ribosome-nascent chain complex
Ribosome-nascent chain complex (RNC)
Post-translational modification (functional groups ·
peptides · structural changes)
Gene regulatory network
Branches of life science and biology
Origin of life
BNF: cb11931064z (data)
Molecular and cellu