The recombinant DNA controversy Those who disregard the past are bound to repeat it George Santayana MCB 140 09 21 07 1 Herb Boyer EcoRI Stanley Cohen pSC101 MCB 140 09 21 07 2 J Bacteriol 64 4 557 569 1952 1953 MCB 140 09 21 07 3 http opbs okstate edu melcher MG MGW3 MG331 html Werner Arber 1965 met depleted E coli don t generate modified phage CH3 of phage DNA MCB 140 09 21 07 4 The restriction modification system a bacterial pathway for defence against viruses MCB 140 09 21 07 5 Dan Nathans Ham Smith MCB 140 09 21 07 6 How restriction enzymes became the workhorses of molecular biology A significant breakthrough came in 1970 when the first of two papers from Smith s laboratory described an enzyme endonuclease R that was able to cleave bacteriophage T7 DNA into specific fragments 2 This was the first type II restriction enzyme the sort that now populates our freezers because it recognize specific sequences and also gives rise to very specific cleavage Smith had been looking for an enzyme that might be involved in site specific recombination in Haemophilus influenzae and thought at first that endonuclease R might be his long sought quarry With Tom Kelly he went on to determine the DNA sequence recognized by endonuclease R and reported it as GTY RAC 11 This sequence seemed too short for a recombination enzyme and during correspondence with his close friend Nathans who ran the neighboring laboratory but was away on sabbatical it became clear that this enzyme might have very practical uses for the analysis of DNA R Roberts PNAS 2005 MCB 140 09 21 07 7 Radioautogram of 14C labeled SV40 DNA cleaved with endonuclease R showing 11 distinct fragments Figure 3 from Danna K Nathans D 1971 Proc Natl Acad Sci USA 68 2913 2917 MCB 140 09 21 07 8 plasmids 9 7 MCB 140 09 21 07 9 9 12 1 8 million MCB 140 09 21 07 10 MCB 140 09 21 07 11 SV40 primate polyomavirus 1972 E coli resident of human GI tract Michael Rogers Biohazard 1977 The question quite simply concerned the wisdom of transplanting SV40 genes conceivably coding for tumor production into a bacterium that not only lacks that capacity to begin with but which lives in virtually every human gut on the planet Berg My God people said You cannot put SV40 into E coli I think I was upset by the criticism at first but then I went out and started to talk about the problem with a lot of people I realized that I d been wrong many many times before in predicting the outcome of an experiment and that if I was wrong about my assessment of the risk in this experiment then the consequences were not something that I would want to live with Berg cancelled the experiment MCB 140 09 21 07 12 Science September 1973 letter to the NAS from participants at Gordon conference on nucleic acids We are writing on behalf of a number of scientists to communicate a matter of deep concern We presently have the technical ability to join together covalently DNA molecules from diverse sources This technique could be used for example to combine DNA from animal viruses with bacterial DNA In this way new kinds of hybrid plasmids or viruses with biological activity of unpredictable nature may eventually be created These experiments offer exciting and interesting potential both for advancing knowledge of fundamental biological processes and for alleviation of human health problems Certain such hybrid molecules may prove hazardous to laboratory workers and to the public Although no hazard has yet been established prudence suggests that the potential hazard be seriously considered Maxine Singer Carnegie Dieter S ll Yale MCB 140 09 21 07 13 September 1974 a moratorium New plasmids novel antibiotic resistance markers into E coli Xenopus and Drosophila genomic DNA into E coli Norton Zinder If we had any guts at all we d tell people to not do these experiments MCB 140 09 21 07 14 MCB 140 09 21 07 15 MCB 140 09 21 07 16 Sept 74 Feb 75 Until the potential hazards of such recombinant DNA molecules have been better evaluated or until adequate methods are developed for preventing their spread scientists throughout the world join with the members of this committee in voluntarily deferring the following experiments cloning new antibiotic resistance genes in currently na ve hosts and cloning fragments of oncoviral genomes into bacterial plasmids MCB 140 09 21 07 17 an epidemic of cancer MCB 140 09 21 07 18 Asilomar conference February 1975 Paul Berg David Baltimore Sydney Brenner Mike Bishop Don Brown Ron Davis James Watson Phil Sharp Herb Boyer Joshua Lederberg 150 more people Nature Science NY Times Washington Post Boston Globe SF Chronicle etc MCB 140 09 21 07 19 Left to right Maxine Singer Norton Zinder Sydney Brenner and Paul Berg MCB 140 09 21 07 20 The measures 1 Moratorium lifted 2 Experiments will proceed under two levels of containment biological use weakened host cell and physical the level of which will be gauged by the level of risk presented by the organism created minimal low moderate high 1 2 3 4 Stringent commonsense cleanliness P1 Similar to surgical operating theatre P2 Giant isolator P3 Comparable to what one uses in biological warfare research P4 MCB 140 09 21 07 21 Where to draw the cutoff line idea 1 invertebrates including flies Stringent commonsense cleanliness P1 vertebrates Similar to surgical operating theatre P2 Giant isolator P3 Comparable to what one uses in biological warfare research P4 MCB 140 09 21 07 22 Where to draw the cutoff line idea 2 invertebrates and cold blooded vertebrates including frogs Warm blooded vertebrates Stringent commonsense cleanliness P1 Similar to surgical operating theatre P2 Giant isolator P3 Comparable to what one uses in biological warfare research P4 MCB 140 09 21 07 23 Enter the sceptics Robert Sinsheimer CalTech The dangers of shotgunning evolutionary jumping creation of chimeras with completely unforeseen new properties Prokaryote to eukatyote gene transfer Use of E coli as a host is potentially dangerous MCB 140 09 21 07 24 Genomic library MCB 140 09 21 07 25 RNA splicing removes introns Exons sequences found in a gene s DNA and mature mRNA expressed regions Introns sequences found in DNA but not in mRNA intervening regions Some eukaryotic genes have many introns MCB 140 09 21 07 26 Dystrophin gene underlying Duchenne muscular dystrophy DMD is an extreme example of introns Fig 8 15 MCB 140 09 21 07 27 Spliced Segments at the 5 Terminus of Adenovirus 2 Late mRNA Susan M Berget Claire Moore and Phillip A Sharp Proceedings of
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