1951-1952 Application for research supvort to theRESEARCH COMMITTEE OF THE UNIVERSITY OF WISCONSIN submitted by Joshua Lederberg, Associate Professor of Genetics January le, 1951 TITLE AND REASONS POR SPuNY Bacterial Genotios. The broad objective of this continuing project is an informed insight into the most fumdarental asrects of microbial blology- the hereditery mechanisms. The most incieive tool of genetic investigation, recombination analycis, is being applied te problere of formal, physiolosical, and cyto-rcenetics in ao many bacteria as ean be shown to be technically feasible. RESULTS OBTAINED TO DATE and PROCEIURE for projected study. Ae Cytogenetic comparisons of haploid and diploid Escherichia coli. -revious reports refer to the discovory of exceptional diploid clones of 5 coli, as indicated by thoir genetic behavior. Those cultures sre boine compared cytolorically with the standard haploid forme, and a critical series of photo~&crographs is in preparation. Naploii end diploid oells ean be distinguished by their nuclear structure, although there io considerable variation in the a pvenranco of gach type through the culture cyclic. The diploid nuclei have a more open, dispersod structuwe than the haploid, but it hao net yet been poseible to resolve coumtable chronose:es, or to outline the presumed mitotic cyolo in detail. The work so far has used pri- narily icilied and staine! orsparations. Uowever, there are published reports, which we are attenctins to verify, thet bacterial mmelei can be resolved in living cells by phase contrast aberoscosy. It ic hoped to use this teehnicue together with the deolation of single celle to correlate cytological ani genetic events of Sef resation. In collaboyrtion with Dr... 2. Zelle of the Aterde Enersy Corrdssion, single coll pedigrees have becn used to verify the dinioid character of the exceptional clones. 3e OQuiterossing bacteria. Toe date, work on conetic roconibination in bacterin has concerned rutant derivatives of a tingle ctradin of Ee coli: "K-12", Attempts to find other bacteria which could be crossed, other introgrossively, or with other strains, had beoa we succonsful, owing to the linited number of tests which could be mado. Methodological Inprovements have made 4t possible, hovever, to ccreen larvo aucbero of strains for "crossability"’ with =12. About 5-105 of isolates ef 2. coll fro humcn urine cultures have proved to be intor- 211 intra-fertile. Among the many research nossLbilities thur opened ur is irwmogenetic onalysie, on which we are gibarlding in a preliminary ways 4 related project, snonnored by the Jetdonal Iastitutes of Tealtiny involves reconbinetion in Salmonella spp. This pro ject Le being pughed in parallel with =. col4 studies, ant has beneMittel ciriilarly fron Srprevel rethole. Ce Bactoricidsl nechanions. In this project, the effects of bactord- elds] agente (ultraviolet light; rays} chewleal rutacone and other ehe-dienic) on diplot? cells heve been examined to try to doterndine how bacteria ore Idlled. There has beon no indfention thet lethe! ratations ploy a role, olthough other nore conplex senretic eects have been fouml. The results oo far have not supported a hit or tarcet thoory, oven for radiations, din contradictiao to moct speculations (Aineluling my own) based upon kinetic data only. The hypothesis presented in a vrevieus report, that "killing is possibly tho ouc- aoedine stare in the series diploid-haplodd-death", ieee, that the nucleus cr chromosome ic the wit of inactivation, has been discardod. The analysis 4a continuing; a research grant from the Atoric enercy Carrmdccdon for X-ray aspects 4s vending. Ye Gene enzyme relationships. Further study of the acture of gonetic control of lactase in Es. cold has been impeded, in part, by the fact that the enzyme 1s fermed in strain 1-1" onlr fin celle which have groom in contact with lactose or oneloseur substrates. Tt wac therefore not feasible to study the cenditione of formation of this oncyme in chert term or manonctric exroriments during which growth is precluded. fortuhately, ome of tic new fortile otrains mentioned above preduces the enzyme unier oinpler conlitionc, and ahonld thus previde moro wor’zable material fer the analysis of lactanse- negative vartanrte. Frelirdinary oxporimants cre now in cropresr to charactorice the lactaces of difforent crosaable ctrainac by the tech- nique of electrophoresis on >apor (cooperntion of or. je Bussard, roscorch fellow, Zoclory Xepte)s It is anticipated that inter-ctrain crosses will give information on the renetic control of onzyme cuality, as well as on the formation of the “standard” oacyic. Larpe-secale preparations avi purification of £. ocli lactase are being explored Dy Dre Te Ae Lardy and “re Se Duby at the Encsyne Institute. AL PITAL suas romosted t 1) Sersomoel. Throe (3) Assi -tantships, 12-n0. § 3606 2 Lado (2) Equipnent Repurchase of rescarch microscope 750 with phase contrast accoseorics “ourly help (for dicheashing and tecimter? rce4ic- tance by undorcradunte students) 300 Total 8 Slane ti on- (1} two of those acelstantships are for renow1: Nios Ethelyn Lively and = Mioe Vhyliic “ried. These asaintarte have carried out their dutics vory satio- fuctorily to date, and would bo difficult te reinco. The third ascistentship is to Bo filled by c candidcte to be selected fron current applications. Unless ur ently required by the best cualified candidate, tiie ascistantclin will be revised to a 1l0-month appointrent. {2} This microncops, originally costing 31200, is in uso in conection with a emtract with the Chenien] cerns on related projocts. This contract will expire durine 1091-52, at which tine title to the microscope will revert. Tho repurchrse price will be subject to nerothation, ani rouy be core or less then the fipure given, althourh the volue cuc ested Ls nrobably a Pedy ono. The type &@f ecubpoout 4. clready difti- eult te obtain, and oucht te be retained.