The recombinant DNA controversySlide 2Slide 3Slide 4The restriction/modification system: a bacterial pathway for defence against virusesSlide 6“How restriction enzymes became the workhorses of molecular biology”Slide 8plasmidsSlide 10Slide 11Slide 12Slide 13September 1974 – a moratorium?Slide 15Slide 16Sept. ’74 – Feb’ 75“an epidemic of cancer”?Asilomar conference, February 1975Slide 20The measuresWhere to draw the cutoff line, idea #1Where to draw the cutoff line, idea #2Enter the scepticsSlide 25RNA splicing removes intronsDystrophin gene underlying Duchenne muscular dystrophy (DMD) is an extreme example of intronsSlide 28Slide 29Slide 30How RNA processing splices out introns and adjoins adjacent exonsSlide 32Slide 33Slide 34Chargaff E, Simring FR. “On the dangers of genetic meddling.” Science. (1976) 192:938Enter the public, part 1“Frankenstein monsters from Harvard Yard”Not only CambridgeEnter the public, part 2Slide 40The severe adverse events that have occurred in the subsequent 30 years of nonstop cloning of all imaginable sorts of DNA into E. coli vectorsA more severe example of the society-genetics interfaceSlide 43Slide 44Technical problemSlide 46Slide 47The two central tenets of LysenkoismLysenko, 19361948: Apocalypse NowCheap populism and demagoguery: a textbook exampleA perspectiveFurther readingA European Perspective on IDWed/FriMCB 140 09-21-07 1The recombinant DNA controversy“Those who disregard the past are bound to repeat it”George SantayanaMCB 140 09-21-07 2Herb Boyer(EcoRI)Stanley Cohen(pSC101)MCB 140 09-21-07 31953J. Bacteriol. 64(4): 557–569 (1952)MCB 140 09-21-07 4http://opbs.okstate.edu/~melcher/MG/MGW3/MG331.htmlWerner Arber (1965): met-depleted E. coli don’t generate “modified” phage CH3 of phage DNA!!MCB 140 09-21-07 5The restriction/modification system:a bacterial pathway for defence against virusesMCB 140 09-21-07 6Dan Nathans Ham SmithMCB 140 09-21-07 7“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 2005MCB 140 09-21-07 8Radioautogram 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, 2913-2917MCB 140 09-21-07 9plasmids9.7MCB 140 09-21-07 109.121.8 millionMCB 140 09-21-07 11MCB 140 09-21-07 12SV40(primate polyomavirus)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.+= ?1972: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 13Science, 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 Söll (Yale)MCB 140 09-21-07 14September 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 15MCB 140 09-21-07 16MCB 140 09-21-07 17Sept. ’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 naïve hosts and cloning fragments of oncoviral genomes into bacterial plasmids.MCB 140 09-21-07 18“an epidemic of cancer”?MCB 140 09-21-07 19Asilomar conference, February 1975Paul 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, etcMCB 140 09-21-07 20(Left to right) Maxine Singer, Norton Zinder, Sydney Brenner, and Paul BergMCB 140 09-21-07 21The measures1. Moratorium lifted.2. Experiments will proceed under two levels of containment, biological (use weakened host cell), and
View Full Document
Unlocking...