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.
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, assuming that no
post-transcriptional regulation occurs and that the levels of mRNA reflect proportional levels of the corresponding protein being produced.
During 1962–1964, through the use of conditional lethal mutants of a bacterial virus, fundamental advances were made in our understanding of the functions and interactions
of the proteins employed in the machinery of DNA replication, DNA repair, DNA recombination, and in the assembly of molecular structures.
 Molecular biology also plays a critical role in the understanding of structures, functions, and internal controls within individual cells, all of which can be used
to efficiently target new drugs, diagnose disease, and better understand cell physiology.
 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.
 Molecular biology is not simply the study of biological molecules and their interactions; rather, it is also a collection of techniques developed since the field’s genesis
which have enabled scientists to learn about molecular processes.
The basic principle is that DNA fragments can be separated by applying an electric current across the gel – because the DNA backbone contains negatively charged phosphate
groups, the DNA will migrate through the agarose gel towards the positive end of the current.
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.
 Techniques of molecular biology For more extensive list on protein methods, see protein methods.
Analogous methods to western blotting can be used to directly stain specific proteins in live cells or tissue sections.
 PCR is a reaction which amplifies small quantities of DNA, and it is used in many applications across scientific disciplines.
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 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.
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
 Western blots can be used to determine the size of isolated proteins, as well as to quantify their expression.
 Another notable contributor to the DNA model was Phoebus Levene, who proposed the “polynucleotide model” of DNA in 1919 as a result of his biochemical experiments on
Griffith advanced another theory, stating that gene transfer occurring in member of same generation is known as horizontal gene transfer (HGT).
Introducing DNA into eukaryotic cells, such as animal cells, by physical or chemical means is called transfection.
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.
PCR can also be used to determine whether a particular DNA fragment is found in a cDNA library.
Molecular cloning Main article: Molecular cloning Transduction image Molecular cloning is used to isolate and then transfer a DNA sequence of interest into a plasmid
In this experiment, as in most molecular biology techniques, a control must be used to ensure successful experimentation.
 Proteins can also be separated on the basis of size 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.
 Molecular biologists today have access to increasingly affordable sequencing data at increasingly higher depths, facilitating the development of novel genetic manipulation
methods in new non-model organisms.
The membrane is then exposed to a labeled DNA probe that has a complement base sequence to the sequence on the DNA of interest.
The plasmid vector usually has at least 3 distinctive features: an origin of replication, a multiple cloning site (MCS), and a selective marker (usually antibiotic resistance).
Molecular genetics, the study of gene structure and function, has been among the most prominent sub-fields of molecular biology since the early 2000s.
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.
Likewise, CRISPR-Cas9 gene editing experiments can now be conceived and implemented by individuals for under $10,000 in novel organisms, which will drive the development of
industrial and medical applications  Relationship to other biological sciences The following list describes a viewpoint on the interdisciplinary relationships between molecular biology and other related fields.
Transduction is a process in which the bacterial DNA carry the fragment of bacteriophages and pass it on the next generation.
A variation of this technique allows the gene expression of an organism at a particular stage in development to be qualified (expression profiling).
PCR has many applications, including the study of gene expression, the detection of pathogenic microorganisms, the detection of genetic mutations, and the introduction of
mutations to DNA.
It is one of the most basic tools for determining at what time, and under what conditions, certain genes are expressed in living tissues.
Likewise, synthetic molecular biologists will drive the industrial production of small and macro molecules through the introduction of exogenous metabolic pathways in various
prokaryotic and eukaryotic cell lines.
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.
 DNA coding for a protein of interest is now inside a cell, and the protein can now be expressed.
Watson and Crick described the structure of DNA and conjectured about the implications of this unique structure for possible mechanisms of DNA replication.
 The concentration of protein in the Bradford assay can then be measured using a visible light spectrophotometer, and therefore does not require extensive equipment.
Genetics attempts to predict how mutations, individual genes and genetic interactions can affect the expression of a phenotype While researchers practice techniques specific
to molecular biology, it is common to combine these with methods from genetics and biochemistry.
Antibodies that specifically bind to the protein of interest can then be visualized by a variety of techniques, including colored products, chemiluminescence, or autoradiography.
This research then lead to finding DNA material in other microorganisms, plants and animals.
The Bradford Assay Main article: The Bradford Assay The Bradford Assay is a molecular biology technique which enables the fast, accurate quantitation of protein molecules
utilizing the unique properties of a dye called Coomassie Brilliant Blue G-250.
 Proteins in the assay bind Coomassie blue in about 2 minutes, and the protein-dye complex is stable for about an hour, although it’s recommended that absorbance readings
are taken within 5 to 20 minutes of reaction initiation.
Arrays can also be made with molecules other than 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 DNA molecule to become 1.07 billion molecules in less than two hours.
 Molecular biology was first described as an approach focused on the underpinnings of biological phenomena – uncovering the structures of biological molecules as well as
their interactions, and how these interactions explain observations of classical biology.
This is also a type of horizontal gene transfer.
 Northern blotting Main article: Northern blot Northern blot diagram The northern blot is used to study the presence of specific RNA molecules as relative comparison
among a set of different samples of RNA.
The field of genetics arose as an attempt to understand the molecular mechanisms of genetic inheritance and the structure of a gene.
[‘1. Alberts B, Johnson A, Lewis J, Morgan D, Raff M, Roberts K, Walter P (2014). Molecular Biology of the Cell, Sixth Edition. Garland Science. pp. 1–10. ISBN 978-1-317-56375-4.
2. ^ Jump up to:a b Gannon F (February 2002). “Molecular biology–what’s
in a name?”. EMBO Reports. 3 (2): 101. doi:10.1093/embo-reports/kvf039. PMC 1083977. PMID 11839687.
3. ^ “Molecular biology – Latest research and news | Nature”. www.nature.com. Retrieved 2021-11-07.
4. ^ Jump up to:a b c d e S., Verma, P. (2004).
Cell biology, genetics, molecular biology, evolution and ecology. S Chand and Company. ISBN 81-219-2442-1. OCLC 1045495545.
5. ^ Astbury, W. T. (June 1961). “Molecular Biology or Ultrastructural Biology ?”. Nature. 190 (4781): 1124. Bibcode:1961Natur.190.1124A.
doi:10.1038/1901124a0. ISSN 1476-4687. PMID 13684868. S2CID 4172248.
6. ^ Jump up to:a b Morange, Michel (2016-02-15), “History of Molecular Biology”, eLS, Chichester, UK: John Wiley & Sons, Ltd, pp. 1–8, doi:10.1002/9780470015902.a0003079.pub3,
ISBN 9780470016176, retrieved 2021-11-07
7. ^ “Polymerase Chain Reaction (PCR)”, Definitions, Qeios, 2019-11-26, doi:10.32388/167113, S2CID 94561339, retrieved 2021-11-07
8. ^ “Smithsonian Institution Archives”. siarchives.si.edu. Retrieved 2021-11-07.
Cox, Michael M. (2015-03-16). Molecular biology: principles and practice. Doudna, Jennifer A.,, O’Donnell, Michael (Biochemist) (Second ed.). New York. ISBN 978-1-4641-2614-7. OCLC 905380069.
10. ^ Bello, Elizabeth A.; Schwinn, Debra A. (1996-12-01).
“Molecular Biology and Medicine: A Primer for the Clinician”. Anesthesiology. 85 (6): 1462–1478. doi:10.1097/00000542-199612000-00029. ISSN 0003-3022. PMID 8968195. S2CID 29581630.
11. ^ Morange, Michel (June 2021). A history of biology. ISBN 978-0-691-18878-2.
12. ^ Fields, Stanley (2001-08-28). “The interplay of biology and technology”. Proceedings of the National Academy of Sciences. 98 (18): 10051–10054. doi:10.1073/pnas.191380098. ISSN 0027-8424. PMC 56913. PMID 11517346.
Ellis, T. H. Noel; Hofer, Julie M. I.; Timmerman-Vaughan, Gail M.; Coyne, Clarice J.; Hellens, Roger P. (2011-11-01). “Mendel, 150 years on”. Trends in Plant Science. 16 (11): 590–596. doi:10.1016/j.tplants.2011.06.006. ISSN 1360-1385. PMID 21775188.
“12.3C: Mendel’s Law of Segregation”. Biology LibreTexts. 2018-07-12. Retrieved 2021-11-18.
15. ^ “Mendelian Inheritance”. Genome.gov. Retrieved 2021-11-18.
16. ^ Jump up to:a b “Discovery of DNA Double Helix: Watson and Crick | Learn Science
at Scitable”. www.nature.com. Retrieved 2021-11-25.
17. ^ George., Wolf (2003). Friedrich Miescher: the man who discovered DNA. OCLC 907773747.
18. ^ Levene, P.A. (1919). “Structure of Yeast Nucleic Acid”. Journal of Biological Chemistry. 43 (2):
379–382. doi:10.1016/s0021-9258(18)86289-5. ISSN 0021-9258.
19. ^ Chargaff, Erwin (1950). “Chemical specificity of nucleic acids and mechanism of their enzymatic degradation”. Experientia. 6 (6): 201–209. doi:10.1007/bf02173653. ISSN 0014-4754.
PMID 15421335. S2CID 2522535.
20. ^ Jump up to:a b Watson, J. D.; Crick, F. H. C. (April 1953). “Molecular Structure of Nucleic Acids: A Structure for Deoxyribose Nucleic Acid”. Nature. 171 (4356): 737–738. Bibcode:1953Natur.171..737W. doi:10.1038/171737a0.
ISSN 1476-4687. PMID 13054692. S2CID 4253007.
21. ^ Crick, F. H. C.; Barnett, Leslie; Brenner, S.; Watts-Tobin, R. J. (1961). “General Nature of the Genetic Code for Proteins”. Nature. Springer Science and Business Media LLC. 192 (4809): 1227–1232.
Bibcode:1961Natur.192.1227C. doi:10.1038/1921227a0. ISSN 0028-0836. PMID 13882203. S2CID 4276146.
22. ^ Epstein, R. H.; Bolle, A.; Steinberg, C. M.; Kellenberger, E.; Boy de la Tour, E.; et al. (1963-01-01). “Physiological Studies of Conditional
Lethal Mutants of Bacteriophage T4D”. Cold Spring Harbor Symposia on Quantitative Biology. Cold Spring Harbor Laboratory. 28: 375–394. doi:10.1101/sqb.1963.028.01.053. ISSN 0091-7451.
23. ^ Mojiri, Soheil; Isbaner, Sebastian; Mühle, Steffen; Jang,
Hongje; Bae, Albert Johann; Gregor, Ingo; Gholami, Azam; Gholami, Azam; Enderlein, Jörg (2021-06-01). “Rapid multi-plane phase-contrast microscopy reveals torsional dynamics in flagellar motion”. Biomedical Optics Express. 12 (6): 3169–3180. doi:10.1364/BOE.419099.
ISSN 2156-7085. PMC 8221972. PMID 34221652.
24. ^ van Warmerdam, T. “Molecular Biology Laboratory Resource”. Yourbiohelper.com.
25. ^ van Warmerdam, T. “Molecular biology laboratory resource”. Yourbiohelper.com.
26. ^ Lodish H, Berk A, Zipursky
SL, Matsudaira P, Baltimore D, Darnell J (2000). Molecular cell biology (4th ed.). New York: Scientific American Books. ISBN 978-0-7167-3136-8.
27. ^ Berg, Jeremy (2002). Biochemistry. Tymoczko, John L.; Stryer, Lubert (5th ed.). New York: W.H.
Freeman. ISBN 0-7167-3051-0. OCLC 48055706.
28. ^ Reference, Genetics Home. “Help Me Understand Genetics”. Genetics Home Reference. Retrieved 31 December 2016.
29. ^ Jump up to:a b Tian J, ed. (2013). Molecular Imaging: Fundamentals and Applications.
Springer-Verlag Berlin & Heidelberg GmbH & Co. K. p. 542. ISBN 9783642343032. Retrieved 2019-07-08.
30. ^ “Foundations of Molecular Cloning – Past, Present and Future | NEB”. www.neb.com. Retrieved 2021-11-25.
31. ^ “Foundations of Molecular Cloning
– Past, Present and Future | NEB”. www.neb.com. Retrieved 2021-11-04.
32. ^ Alberts B, Johnson A, Lewis J, Raff M, Roberts K, Walter P. Isolating, Cloning, and Sequencing DNA. Retrieved 31 December 2016.
33. ^ Lessard, Juliane C. (1 January 2013).
“Molecular cloning”. Laboratory Methods in Enzymology: DNA. Methods in Enzymology. Vol. 529. pp. 85–98. doi:10.1016/B978-0-12-418687-3.00007-0. ISBN 978-0-12-418687-3. ISSN 1557-7988. PMID 24011038.
34. ^ Kokate C, Jalalpure SS, Hurakadle PJ (2016).
Textbook of Pharmaceutical Biotechnology. Expression Cloning. Elsevier. p. 125. ISBN 9788131239872. Retrieved 2019-07-08.
35. ^ Lenstra, J. A. (July 1995). “The applications of the polymerase chain reaction in the life sciences”. Cellular and Molecular
Biology (Noisy-Le-Grand, France). 41 (5): 603–614. ISSN 0145-5680. PMID 7580841.
36. ^ “Polymerase Chain Reaction (PCR)”. National Center for Biotechnology Information. U.S. National Library of Medicine. Retrieved 31 December 2016.
37. ^ “Polymerase
Chain Reaction (PCR) Fact Sheet”. National Human Genome Research Institute (NHGRI). Retrieved 31 December 2016.
38. ^ Jump up to:a b Lee, Pei Yun; Costumbrado, John; Hsu, Chih-Yuan; Kim, Yong Hoon (2012-04-20). “Agarose Gel Electrophoresis for the
Separation of DNA Fragments”. Journal of Visualized Experiments (62): 3923. doi:10.3791/3923. ISSN 1940-087X. PMC 4846332. PMID 22546956.
39. ^ Lee PY, Costumbrado J, Hsu CY, Kim YH (April 2012). “Agarose gel electrophoresis for the separation of
DNA fragments”. Journal of Visualized Experiments (62). doi:10.3791/3923. PMC 4846332. PMID 22546956.
40. ^ Jump up to:a b c d e f Bradford, Marion M. (1976-05-07). “A rapid and sensitive method for the quantitation of microgram quantities of protein
utilizing the principle of protein-dye binding”. Analytical Biochemistry. 72 (1): 248–254. doi:10.1016/0003-2697(76)90527-3. ISSN 0003-2697. PMID 942051.
41. ^ Jump up to:a b c “Protein determination by the Bradford method”. www.ruf.rice.edu. Retrieved
42. ^ Thomas PS (September 1980). “Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose”. Proceedings of the National Academy of Sciences of the United States of America. 77 (9): 5201–5. Bibcode:1980PNAS…77.5201T.
doi:10.1073/pnas.77.9.5201. PMC 350025. PMID 6159641.
43. ^ Brown T (May 2001). “Southern blotting”. Current Protocols in Immunology. Chapter 10: Unit 10.6A. doi:10.1002/0471142735.im1006as06. ISBN 978-0-471-14273-7. PMID 18432697. S2CID 20686993.
Josefsen K, Nielsen H (2011). Nielsen H (ed.). RNA methods and protocols. Methods in Molecular Biology. Vol. 703. New York: Humana Press. pp. 87–105. doi:10.1007/978-1-59745-248-9_7. ISBN 978-1-59745-248-9. PMID 21125485.
45. ^ He SL, Green R (1
January 2013). “Northern blotting”. Methods in Enzymology. 530: 75–87. doi:10.1016/B978-0-12-420037-1.00003-8. ISBN 978-0-12-420037-1. PMC 4287216. PMID 24034315.
46. ^ Jump up to:a b Mahmood T, Yang PC (September 2012). “Western blot: technique,
theory, and trouble shooting”. North American Journal of Medical Sciences. 4 (9): 429–34. doi:10.4103/1947-2714.100998. PMC 3456489. PMID 23050259.
47. ^ “Western blot | Learn Science at Scitable”. www.nature.com. Retrieved 2021-11-25.
48. ^ Kurien
BT, Scofield RH (April 2006). “Western blotting”. Methods. 38 (4): 283–93. doi:10.1016/j.ymeth.2005.11.007. PMID 16483794. – via ScienceDirect (Subscription may be required or content may be available in libraries.)
49. ^ Thomas S, Thirumalapura
N, Crossley EC, Ismail N, Walker DH (June 2009). “Antigenic protein modifications in Ehrlichia”. Parasite Immunology. 31 (6): 296–303. doi:10.1111/j.1365-3024.2009.01099.x. PMC 2731653. PMID 19493209.
50. ^ “Microarrays”. National Center for Biotechnology
Information. U.S. National Library of Medicine. Retrieved 31 December 2016.
51. ^ Bumgarner R (January 2013). Frederick M. Ausubel, et al. (eds.). “Overview of DNA microarrays: types, applications, and their future”. Current Protocols in Molecular
Biology. Chapter 22: Unit 22.1. doi:10.1002/0471142727.mb2201s101. ISBN 978-0-471-14272-0. PMC 4011503. PMID 23288464.
52. ^ Govindarajan R, Duraiyan J, Kaliyappan K, Palanisamy M (August 2012). “Microarray and its applications”. Journal of Pharmacy
& Bioallied Sciences. 4 (Suppl 2): S310-2. doi:10.4103/0975-7406.100283. PMC 3467903. PMID 23066278.
53. ^ Tarca AL, Romero R, Draghici S (August 2006). “Analysis of microarray experiments of gene expression profiling”. American Journal of Obstetrics
and Gynecology. 195 (2): 373–88. doi:10.1016/j.ajog.2006.07.001. PMC 2435252. PMID 16890548.
54. ^ Cheng L, Zhang DY, eds. (2008). Molecular genetic pathology. Totowa, NJ: Humana. p. 96. ISBN 978-1-59745-405-6. Retrieved 31 December 2016.
Leonard DG (2016). Molecular Pathology in Clinical Practice. Springer. p. 31. ISBN 978-3-319-19674-9. Retrieved 31 December 2016.
56. ^ Tian J, ed. (2013). Molecular Imaging: Fundamentals and Applications. Springer-Verlag Berlin & Heidelberg GmbH
& Co.K. pp. 550, 552. ISBN 9783642343032. Retrieved 2019-07-08.
Photo credit: https://www.flickr.com/photos/orangeaurochs/8614237582/’]