50 Recombinant DNA | Summary

Recombinant DNA

Recombinant DNA is an artificial form of DNA that cannot be found in natural organisms. It is made in the laboratory by joining together genes taken from different sources. This is done by selecting and cutting out a gene at a specific point on a strand of DNA using restriction enzymes which act as molecular scissors. The gene is chosen on the basis of its ability to code for or alter different traits in another organism. It is inserted into a circular piece of bacterial DNA, called a plasmid, or a bacterial virus, called a phage, and then put into a host organism, such as the bacteria Escherichia-Coli, for replication by its cell machinery.

The vial shown in the photograph contains human insulin, one of the first therapeutic proteins that was genetically cloned. The drug is used to treat diabetes. Credit: Wellcome Library, London.

Definition

Recombinant DNA is a form of DNA constructed in the laboratory. It is generated by transferring selected pieces of DNA from one organism to another.

Importance

Genetic engineering is used for many different purposes in research, medicine, agriculture and industry. The technology is important because it enables the creation of multiple copies of genes and the insertion of foreign genes into other organisms to give them new traits, such as antibiotic resistance or a new colour. One of the first ways in which the technology was deployed was to re-engineer microbial cells to produce foreign proteins. This facilitated the manufacture of human proteins on an unprecedented scale at minimum cost, thereby opening the way to study the function of proteins in greater detail and to their therapeutic use. By 2001 over 80 recombinant DNA based products had been approved for treating disease and for vaccination and a further 350 recombinant DNA-based drugs were being tested for safety and efficacy. The technology is also an important tool in agriculture, being used to improve plants' resistance to pests and increase crop yields.

Discovery

While the structure of DNA was first determined in 1953, it was to take another two decades before scientists had the means to generate recombinant DNA. This was aided by firstly the realisation in the 1950s that plasmids, small mobile pieces of DNA, could replicate in huge quantities independently of chromosomal bacteria DNA and that they could transfer genetic information. It was this process that gave host bacteria the capacity to inherit new genes and therefore new functions such as resistance to antibiotics. Another important tool for creating recombinant DNA was the discovery in the 1960s by the Swiss microbiologist Werner Arber and American biochemist Stuart Linn that bacteria could protect themselves from attack by viruses the production of endonucleases, known as restriction enzymes, which could seek out a single DNA sequence in a virus and cut it precisely in one place. This process prevented the replication of viruses and hence the death of virally infected bacteria. The first restriction enzyme, Escscherichia coli K, was isolated and purified in 1968 by Matthew Meselson and Robert Yuan at Harvard University. Two years later Hamilton O Smith, Thomas Kelly and Kent Welcox at Johns Hopkins University isolated and characterised the first site-specific restriction enzyme, later named HindII. This was demonstrated by Daniel Nathans to be a useful tool for cutting and pasting specific DNA segments. The first protocol for creating recombinant DNA was put forward in the early 1970s by Peter Lobban and Armin Dale Kaiser at Stanford University Medical School. In 1971 Paul Berg, attached to Stanford University, demonstrated the feasibility of splicing and recombining genes for the first time. Two years later, Stanley Cohen and Herbert Boyer, based respectively at Stanford University and University of California at San Francisco, successfully inserted recombinant DNA into bacteria for replication.

Application

Gene cloning has a diverse range of applications. Where it has proven particularly useful has been in mapping out the human genome, the creation of transgenic animals, and the development of insect-resistant crops. It is also pivotal to genetic tests carried out in forensic science and archaeology as well as in tests for determining hereditary disease and paternity. The technology also forms the backbone of hepatitis and human immunodeficiency virus (HIV) diagnostic tests. Recombinant DNA technology has also proven important to the production of vaccines and protein therapies such as human insulin, interferon and human growth hormone. It is also used to produce clotting factors for treating haemophilia and in the development of gene therapy.

Recombinant DNA: timeline of key events

Date Event People Places
December 14, 1914Solomon Spiegelman was born in Brooklyn, NY, USASpiegelmanUniversity of Minnesota
June 30, 1926Paul Berg was born in New York NY, USABergStanford University
June 3, 1929Werner Arber was born in Granichen, SwitzerlandArberUniversity of Geneva
April 26, 1932Michael Smith was born in Blackpool, United KingdomSmithUniversity of British Columbia
June 30, 1935Stanley Norman Cohen was born in Perth Amboy, NJ, USACohenStanford University
1941Term 'genetic engineering' first coinedJost
1952First observation of the modification of viruses by bacteriaLuria, HumanUniversity of Illinois
December 1955First discovery of the enzyme DNA polymeraseKornberg, Bessman, Simms, LehmanWashington University in St. Louis
October 1957First synthesis of DNA in a test tubeKornbergWashington University in St. Louis
1962Concept of restriction and modification enzymes bornArber, DussoixUniversity of Geneva
1965Werner Arber predicts restriction enzymes could be used as a labortory tool to cleave DNAArberUniversity of Geneva
1968 - 1970Restriction enzymes found to act as chemical knives to cut DNASmith, NathansUniversity of Geneva, University of California in Berkeley, Johns Hopkins University
1968Paul Berg started experiments to generate recombinant DNA moleculesBergStanford University
1969New idea for generating recombinant DNA conceivedLobhanStanford University
1970First complete gene synthesised KhoranaUniversity of Wisconsin
July 1970First restriction enzyme isolated and characterisedSmith, WilcoxJohns Hopkins University
1971First plasmid bacterial cloning vector constructedBerg, Mertz, JacksonStanford University
June 1971First time potential biohazards of recombinant DNA were raisedMertz, Berg, PollackStanford University
December 1971First experiments published demonstrating the use of restriction enzymes to cut DNADanna, NathansJohns Hopkins University
1972First recombinant DNA createdBerg, MertzStanford University Medical School
September 1972 - September 1972First time possible biohazards of recombinant DNA technology publicly discussedZinder
October 1972First paper published on generating recombinant DNABerg, Jackson, SymonsStanford University
November 1, 1972Nature editorial voiced concern about generating recombinant DNABerg, Jackson, SymonsStanford University
1973Recombinant DNA produced in bacteriaCohen, BoyerStanford University Medical School, University of California San Francisco
1974Regulation begins for recombinant genetic research
July 1974First concerns about potential biohazards of recombinant DNA publishedBerg, Baltimore, Boyer, Cohen
1975Temporary moratorium on genetic engineeringBerg
April 1976Genentech foundedSwanson, BoyerGenentech Inc
1977Human growth hormone genetically engineered
1978Human insulin produced in E-coliGenentech
October 1978Nobel Prize for discovery and understanding of restriction enzymesArber, Nathans, SmithJohns Hopkins University, University of Geneva
1980Genetic engineering recognised for patenting
1980First patent awarded for gene cloningCohen, BoyerStanford University Medical School
1980Cesar Milstein proposed the use of recombinant DNA to improve monoclonal antibodiesMilsteinLaboratory of Molecular Biology
September 1980First transgenic mice made with recombinant DNA announced Barbosa, Gordon, Plotkin, Ruddle, ScangosYale University
1981First genetically-engineered plant reported
1981First genetically cloned mice
October 1982First recombinant DNA based drug approvedGenentech Inc
January 20, 1983Solomon Spiegelman diedSpiegelmanUniversity of Minnesota
1984First chimeric monoclonal antibodies developed which lays foundation for safer and more effective monoclonal antibody therapeuticsNeuberger, Rabbitts, Morrison, Oi, Herzenberg, Boulianne, Schulman, HozumiLaboratory of Molecular Biology, Stanford Univerity Medical School
May 1986First humanised monoclonal antibody createdDear, Foote, Jones, Neuberger, WinterLaboratory of Molecular Biology
June 1986Interferon approved for treating hairy cell leukaemia
June 1986First recombinant vaccine licensedMerck & Co
1988Campath-1H is created - the first clinically useful humanised monoclonal antibody.Winter, Waldmann, Reichmann, ClarkCambridge University, Laboratory of Molecular Biology
December 1993Recombinant human deoxyribonuclease approved by FDA for cystic fibrosisGenentech
1994First chimeric monoclonal antibody therapeutic approved for marketCollerCentocor, State University of New York
December 1997First humanised monoclonal antibody approved for marketQueenProtein Design Labs, Roche
November 16, 1999Daniel Nathans diedNathans Johns Hopkins University
October 4, 2000Michael Smith diedUniversity of British Columbia
March 2014Promising results announced from trial conducted with HIV patients

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