Molecular biology ( /məˈlɛkjʊlər/) is the branch of biology that deals with the molecular basis of biological activity. This field overlaps with other areas of biology and chemistry, particularly genetics and biochemistry. Molecular biology chiefly concerns itself with understanding the interactions between the various systems of a cell, including the interactions between the different types of DNA, RNA and protein biosynthesis as well as learning how these interactions are regulated.

Writing in Nature in 1961, 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[link]

Schematic relationship between biochemistry, genetics, and molecular biology

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 above is a schematic that depicts one possible view of the relationship between the fields:

• Biochemistry is the study of the chemical substances and vital processes occurring in living 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. Often this can 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 "knock-out" 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 an oversimplified picture of molecular biology, still provides a good starting point for understanding the field. This picture, however, is undergoing revision in light of emerging novel roles for RNA.

Much of the work in molecular biology is quantitative, and recently much work has been done at the interface of molecular biology and computer science in bioinformatics and computational biology. As of the early 2000s, the study of gene structure and function, molecular genetics, has been among the most prominent sub-field of molecular biology.

Increasingly many other loops of biology focus on molecules, either directly studying their interactions in their own right such as in cell biology and developmental biology, or indirectly, where the techniques of molecular biology 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 biophysics.

Techniques of molecular biology[link]

Since the late 1950s and early 1960s, molecular biologists have learned to characterize, isolate, and manipulate the molecular components of cells and organisms. These components include DNA, the repository of genetic information; RNA, a close relative of DNA whose functions range from serving as a temporary working copy of DNA to actual structural and enzymatic functions as well as a functional and structural part of the translational apparatus; and proteins, the major structural and enzymatic type of molecule in cells.

Expression cloning[link]

One of the most basic techniques of molecular biology to study protein function is expression cloning. In this technique, DNA coding for a protein of interest is cloned (using PCR and/or restriction enzymes) into a plasmid (known as an expression vector). A vector has 3 distinctive features: an origin of replication, a multiple cloning site (MCS), and a selective marker (usually antibiotic resistance). The origin of replication will have promoter regions upstream the replication/transcription start site.

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. DNA can also be introduced into eukaryotic cells using viruses or bacteria as carriers, the latter is sometimes called bactofection and in particular uses Agrobacterium tumefaciens. The plasmid may be integrated into the genome, resulting in a stable transfection, or may remain independent of the genome, called transient transfection.

In either case, 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 (PCR)[link]

The polymerase chain reaction is an extremely versatile technique for copying DNA. In brief, PCR allows a single DNA sequence to be copied (millions of times), or altered in predetermined ways. For example, PCR can be used to introduce restriction enzyme sites, or to mutate (change) particular bases of DNA, the latter is a method referred to as "Quick change". PCR can also be used to determine whether a particular DNA fragment is found in a cDNA library. PCR has many variations, like reverse transcription PCR (RT-PCR) for amplification of RNA, and, more recently, real-time PCR (QPCR) which allow for quantitative measurement of DNA or RNA molecules.

Gel electrophoresis[link]

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. In agarose gel electrophoresis, DNA and RNA can be separated on the basis of size by running the DNA through an 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[link]

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.^[2] Further combinations of these techniques produced such terms as southwesterns (protein-DNA hybridizations), northwesterns (to detect protein-RNA interactions) and farwesterns (protein-protein interactions), all of which are presently found in the literature.

Southern blotting[link]

Named after its inventor, biologist Edwin Southern, the Southern blot is a method for probing for the presence of a specific DNA sequence within a DNA sample. DNA samples before or after restriction enzyme 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 DNA probe that has a complement base sequence to the sequence on the DNA of interest. Most original protocols used radioactive labels, however non-radioactive alternatives are now available. Southern blotting is less commonly used in laboratory science due to the capacity of other techniques, such as PCR, to detect specific 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.

Northern blotting[link]

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 denaturing RNA gel electrophoresis, and a blot. In this process RNA is 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 RNA is 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.

Western blotting[link]

Antibodies to most proteins can be created by injecting small amounts of the protein into an animal such as a mouse, rabbit, sheep, or donkey (polyclonal antibodies) or produced in cell culture (monoclonal antibodies). These antibodies can be used for a variety of analytical and preparative techniques.

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 (sodium dodecyl sulfate polyacrylamide gel electrophoresis). The proteins in the gel are then transferred to a 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. However, these immunostaining methods, such as FISH, are used more often in cell biology research.

Eastern blotting[link]

Eastern blotting technique is to detect post-translational modification of proteins.^[3] Proteins blotted on to the PVDF or nitrocellulose membrane are probed for modifications using specific substrates.

Arrays[link]

A DNA array is a collection of spots attached to a solid support such as a microscope slide where each spot contains one or more single-stranded 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 (similar to Southern blotting). 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 the RNA in a tissue is isolated and converted to labeled cDNA. This cDNA 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. For instance, the common baker's yeast, Saccharomyces cerevisiae, contains about 7000 genes; with a microarray, one can measure qualitatively how each gene is expressed, and how that expression changes, for example, with a change in temperature. 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. For example, an antibody array can be used to determine what proteins or bacteria are present in a blood sample.

Allele Specific Oligonucleotide[link]

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 target 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 successful experimentation. The Illumina Methylation Assay is an example of a method that takes advantage of the ASO technique to measure one base pair differences in sequence.

Antiquated technologies[link]

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 size of 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.

History[link]

While molecular biology was established in the 1930s, the term was coined by Warren Weaver in 1938. Warren 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. He therefore channeled significant amounts of (Rockefeller Institute) money into biological fields.

Clinical significance[link]

Clinical research and medical therapies arising from molecular biology are partly covered under gene therapy^{[citation needed]}

This section requires expansion.

The use of molecular biology or molecular cell biology approaches in medicine is now called Molecular Medicine.

See also[link]

Molecular and Cellular Biology portal

References[link]

• Cohen, S.N., Chang, A.C.Y., Boyer, H. & Heling, R.B. Construction of biologically functional bacterial plasmids in vitro. Proc. Natl. Acad. Sci. 70, 3240 – 3244 (1973).
• Rodgers, M. The Pandora's box congress. Rolling Stone 189, 37 – 77 (1975).

Further reading[link]

• Keith Roberts, Martin Raff, Bruce Alberts, Peter Walter, Julian Lewis and Alexander Johnson, Molecular Biology of the Cell
• 4th Edition, Routledge, March, 2002, hardcover, 1616 pages, 7.6 pounds, ISBN 0-8153-3218-1
• 3rd Edition, Garland, 1994, ISBN 0-8153-1620-8
• 2nd Edition, Garland, 1989, ISBN 0-8240-3695-6

External links[link]

• Biochemistry and Molecular Biology at the Open Directory Project

Drew Berry (born 1970 in USA) is a biomedical animator at the Walter and Eliza Hall Institute of Medical Research in Melbourne, Australia. His scientifically accurate and aesthetically rich visualizations are elucidating cellular and molecular processes for a wide range of audiences. His animations have been shown in exhibitions, multimedia programs and television shows, and have received international recognition including an Emmy (2005) and a BAFTA Award (2004).

Education[link]

Berry received a Bachelor of Science (1993) and Master of Science (1995) degrees from the University of Melbourne, and received training in cell biology as well as in light microscopy and electron microscopy.

Career[link]

Since 1995, Berry has been a biomedical animator at the Walter and Eliza Hall Institute of Medical Research.^[1] His 3D and 4D animations have focussed on explaining cellular and molecular processes relevant to research conducted at the institute, in fields including molecular biology, malaria, cell death, cancer biology, hematology and immunology.^[2]

Berry's animations have received many awards and commendations. His animations which formed part of Harold Varmus' 'Genes and Jazz' presentation were described by The New Yorker as "astonishly beautiful".^[3] In 2009 American Scientist stated "The admirers of Drew Berry... talk about him the way Cellini talked about Michelangelo."^[4] In 2010, the New York Times claimed “If there is a Steven Spielberg of molecular animation, it is probably Drew Berry”^[5]

After collaborating with Berry for the Biophilia animated music app, the musician Björk described Berry as “someone who has made scientifically correct animation of DNA… on this project, he has crossed the line beautifully into the artistic realm where hehas animated gorgeous DNA but added some poetic licence... he truly has brought magic to our insides, and shows us that we don’t have to look far for the miracle of nature, it is right inside us!” ^[6]

Awards and Recognition[link]

• 2004- BAFTA award for DNA Interactive DVD, UK 2004^[7]
• 2005- Emmy Award for DNA documentary series^[8]
• 2005- Maya Master Award
• 2006- National Science Foundation Science and Engineering Visualization Challenge first place, noninteractive multimedia^[9]
• 2008- Nature Niche Prize^[10]
• 2009- Seed Magazine 'Revolutionary Minds'^[11]
• 2010- MacArthur Fellowship^[12]

Highlight Exhibitions and Productions[link]

• 2003- SIGGRAPH Electronic Theater^[13]
• 2003- “DNA” project Channel 4, PBS and American Museum of Natural History; Five episode “DNA” documentary series, “DNA: The Secret of Life” museum film, “DNAi” DVD, DNAi.org online education portal [13]
• 2003- “oZone” Cinema of Tomorrow Experimental Digital Media Art Festival, Pompidou Centre Paris, France^[14]
• 2003- International Genetics Congress 2003 opening ceremony performance, Melbourne, Australia^[15]
• 2003-04- Australian Centre for the Moving Image Federation Square ‘Transfigure’ exhibition, Australia^[16]
• 2004-05- Museum of Modern Art (New York) ‘Premieres’ exhibition^[17]
• 2004-05- Queensland Art Gallery “The Nature Machine” exhibition, Brisbane, Australia^[18]
• 2004-2006- National Academy of Sciences Koshland Science Museum, USA^[19]
• 2005- Moving Image Centre “Transfigure” exhibition, Auckland, New Zealand
• 2005 Visual effects for Doctor Who episode "The End of the World"
• 2006- Rose Center for Earth and Space, American Museum of Natural History, New York
• 2006- Shanghai Zendai Museum of Modern Art ‘Strange Attractors’ exhibition, China^[20]
• 2006-07- Forum Kultur und Wirtschaft Duesseldorf, Museum of Design, exhibition on nanotechnology, Germany^[21]
• 2008- 20th Century Fox ‘The Day The Earth Stood Still’ visual effects, USA^[22]
• 2008-09- The Royal Institute of Great Britain installation, UK
• 2008-09- ‘Genes and Jazz’ at the Guggenheim with Harold Varmus^[23]
• 2009- ‘The Cell’ [14] BBC documentary series with Dr Adam Rutherford, UK
• 2009- Museum of Natural History Stuttgart ‘200th birthday of Charles Darwin’ exhibition, Germany^[24]
• 2009-10- University of Geneva’s ‘Genome Dome’ exhibition, Switzerland
• 2010- Imagine Science Film Festival, New York^[25]
• 2011- TEDxCaltech^[26]
• 2011- TEDxSydney^[27]
• 2011- Created animations to accompany the track Hollow, in Björk's Biophilia album iPad app ^[28]
• 2011- Rochester Institute of Technology "Visionaries in Motion" speaker series^[29]
• 2011- Cold Spring Harbor Laboratory's DNAi.org Spliceosome visualization
• 2012- Animations for E.O.Wilson's Life on Earth digital text book ^[30]

References[link]

1. ^ Drew Berry by Colin Martin, The Lancet, Volume 377, Issue 9769, Page 895 [1] 12 March 2011
2. ^ WEHI.TV Animations
3. ^ Swing Science by Paul Goldberger, The New Yorker [2] December 1, 2008
4. ^ American Scientist, Science Observer: Molecular Movie Stars [3] September–October 2009 Volume 97, Number 5 Page: 382
5. ^ Where Cinema and Biology Meet [4] 15 November 2010.
6. ^ Drew Berry's Bio-Animations Dazed Digital Dazed & Confused August 2011 Issue
7. ^ Twisted tale of the double helix [5] 29 February 2004
8. ^ Animations for the multi-media project, DNA, author Drew Berry, wins Emmy Award [6] 23 September 2005.
9. ^ 2006 Science and Engineering Visualization Challenge winners announced [7] 21 September 2006
10. ^ Niche Prize winners announced [8] 16 July 2008
11. ^ Revolutionary Minds: The Interpreters: Drew Berry- illuminated manuscripts [9] April 2009.
12. ^ 2010 MacArthur Fellows: Drew Berry
13. ^ SIGGRAPH 2003, San Diego, Electronic Theater
14. ^ About WEHI.TV
15. ^ About WEHI.TV
16. ^ Drew Berry- Body Code, Australian Centre for the Moving Image [10] 2003
17. ^ MoMA Film Exhibitions: Premieres November 21, 2004–January 31, 2005
18. ^ The Nature Machine Media Kit, Queensland Art Gallery
19. ^ About WEHI.TV
20. ^ Strange Attractors: Exhibiting Artists
21. ^ About WEHI.TV
22. ^ New York Times- 'The Day the Earth Stood Still' production credits
23. ^ In Science and Jazz, Father and Son Find Common Bonds [11] 9 June 2009
24. ^ About WEHI.TV
25. ^ Imagine Science Films: Fighting Infection By Clonal Selection [12] 18 October 2010
26. ^ TEDxCaltech speakers
27. ^ TEDxSydney speakers
28. ^ Drew Berry's Bio-Animations Dazed Digital Dazed & Confused August 2011 Issue
29. ^ "Biomedical Animator Drew Berry to Speak at RIT Dec. 8" RIT University News Dec 1, 2011
30. ^ "E. O. Wilson's Life on Earth: Team"

External links[link]

• WEHI.TV, Walter and Eliza Hall Institute
• Hollow, animation for Björk's Biophilia album, 2011.