Influential ideas are always simple Since natural phenomena need not be simple we master them if at all by formulating simple ideas and exploring their limitations Al Hershey MCB 140 11 3 05 1 MCB 140 11 3 05 2 MCB 140 11 3 05 3 MCB 140 11 3 05 4 Bacterial genetics I MCB 140 11 3 05 5 What are the genes What is the nature of the elements of heredity that Mendel postulated as purely theoretical units Frankly these are questions with which the working geneticist has not much concern himself If the gene is a material unit it is a piece of a chromosome if it is a fictitious unit it must be referred to a definite location in a chromosome Therefore it makes no difference in the actual work in genetics which point of view is taken T H Morgan The Relation of Genetics to Physiology and Medicine Nobel Lecture June 4 1934 MCB 140 11 3 05 6 DNA RNA protein central dogma of molecular biology MCB 140 2 25 05 7 7 17 MCB 140 11 3 05 8 Max Delbr ck 1906 1981 Salvador Luria 1912 1991 Cold Spring Harbor Laboratory Long Island NY Summer 1941 MCB 140 11 3 05 9 Today even the layman thinks of resistant bacteria as originating from mutation but when Luria and Delbr ck first got together conventional bacteriologists were by no means clear that microorganisms could be tought about genetically Many believed that resistance was some kind of adaptation induced in a few of the bacteria in a culture by the exposure to the antibacterial agent Judson p 55 MCB 140 11 3 05 10 The idea smacks of the preMendelian pre Darwinian notion of the inheritance of acquired characteristics Luria damned bacteriology as the last stronghold of Lamarckism Judson p 55 MCB 140 11 3 05 11 Let s all do science in Nevada One Saturday evening Luria went to a faculty dance There watching the fluctuating returns obtained by colleagues gambling on a slot machine he thought of the experiment that would distinguish between resistance induced in bacteria and resistance resulting from previous spontaneous mutation upon which selection acts Judson p 55 MCB 140 11 3 05 12 What Luria perceived was that previous spontaneous mutation would pay out jackpots of resistant bacteria that would fluctuate much more widely in size than those paid out by induction He tried the first experiment on the following morning and wrote off to Delbrueck Delbrueck promptly replied that Luria really ought to go to church Judson p 55 MCB 140 11 3 05 13 7 4 MCB 140 11 3 05 14 What Luria actually did Sample set A 1 Inoculate bacteria into individual cultures 1 bacterium per culture 2 Let it grow up to a large number Sample set B 1 Take an aliquot of bacteria and start a culture which will therefore not be clonal 2 Let them grow up to a large number Expose both to phage and count how many phageresistant colonies per culture are found Ask if there is a difference between these two sample sets MCB 140 11 3 05 15 S Luria M Delbr ck 1943 Mutations of bacteria from virus sensitivity to virus resistance Genetics 28 491 511 If the production of resistance began only at the moment of exposure to phage then it wouldn t matter whether the bacteria came from many individual cultures or one bulk culture When Luria performed the experiment though the twenty separate cultures showed much wider fluctuations from the average number of resistant colonies indicating that a few of the individual tubes contained resistant bacteria from near the beginning of the overnight growth period Judson p 56 MCB 140 11 3 05 16 Brock p 59 MCB 140 11 3 05 17 MCB 140 11 3 05 18 MCB 140 11 3 05 19 Remember Prof Amacher s lecture 8 George Beadle left and Edward Tatum right receiving their Nobel Prizes MCB 140 11 3 05 20 MCB 140 11 3 05 21 7 20 MCB 140 11 3 05 22 Joshua Lederberg MCB 140 11 3 05 23 Conjugation in bacteria Take strain of E coli that is auxotrophic for two distinct nutrients thiamine and leucine Take different strain of E coli that is also auxotrophic for two distinct nutrients but different ones biotine and cysteine Mix the two Ask if ANY NOVEL PHENOTYPES APPEAR MCB 140 11 3 05 24 J Lederberg E Tatum 1946 Novel genotypes in mixed cultures of biochemical mutants of bacteria Cold Spring Harbor Symp Quant Biol 11 113 114 14 11 MCB 140 11 3 05 25 Paramecium MCB 140 11 3 05 26 B H and A H In the pre Hayes period mating in bacteria was envisioned as a conventional sex process perhaps modified by aspects of relative sexuality but nevertheless a standard haploid diploid meiosis mechanism After Hayes it was known that bacteria were not just small cells but constituted a completely different kind of cell The terms prokaryote and eukaryote were not introduced until 1962 Brock p 87 MCB 140 11 3 05 27 William Hayes MCB 140 11 3 05 28 Selman Waksman streptomycin MCB 140 11 3 05 29 Hayes expt Take strain A which is streptomycin resistant and auxotrophic for biotin and methionine Take strain B which is streptomycin sensitive and auxotrophic for threonine and leucine Mix the two on a minimalmedium plate containing streptomycin Wait and see MCB 140 11 3 05 30 Rich husband poor wife is not the same as poor husband rich wife Cross 1 Strain A StrR B M Strain B StrS L T Result streptomycin completely inhibits prototroph formation i e appearance of B M L T bacteria if added before conjugation is complete Cross 2 Strain A StrS B M Strain B StrR L T Result streptomycin has no effect whatsoever You can add it all you want at any time and prototrophs will still form MCB 140 11 3 05 31 I discussed these results with Denny Mitchison and I think it was he who first suggested that one of the parents A might be acting as a gene donor and the other B as a recipient It was from this experiment that the concept of asymmetry in bacterial sexuality arose Parent B was the recipient or female the continued viability of which was essential for the whole process of recombination and segregation while the A donor or male cell was dispensable once genetic transfer had been effected W Hayes 1952 Recombination in Bact coli K 12 unidirectional transfer of genetic material Nature 169 118 Brock p 89 MCB 140 11 3 05 32 MCB 140 11 3 05 33 14 12 MCB 140 11 3 05 34 14 12 MCB 140 11 3 05 35 14 13 MCB 140 11 3 05 36 14 14 MCB 140 11 3 05 37 E Wollman F Jacob 1955 Sur le mecanisme du transfert de materiel genetique au cours de la recombinaison chez E coli K12 CR Academie Sciences 240 2449 MCB 140 11 3 05 38 MCB 140 11 3 …
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