. li a4P Project: Geneties of Bacteria Joshua Lederberg Asat. Professor of Genetics University of Wisconein Three modes of hereditary transmission are now recognized to occur in bacteria. The first is simply the production of lixe by like in the fission or vegetative growth of bacteri=l cells, The second ia embodied in "sexual" reproduction, where- by two cella onn exchange or recombine their hereditary particles, giving rise to new genotypeer distinct from either parent, The third is infective transmission, whereby » hereditary charecter of one microbial line may be trenemitted vin the medium to another, Of these three modes, the firet two are of course found in nearly all forms of life; the third (apart from the behavior of parasitic viruses) is a new principle which has been firmly established only in recent years (1) through the investigation of transformations of type in pneumococcus, Sexual re- production is an even newer tool in the hands of the bacterial geneticist. The author's recent investigotions of thie phenomenon are reported in the reprints at- teched to this memorandum, The research program which is planned and under way in this laboratory can be categorized under two heedings, No, 1 ie an application of the tool of breeding analyeia in dacteria to a problem in gene action--the relationship between gene matationa and changer in enzymetic epecificity, which is supposed tc represent the primary function of the gene, No, 2 ie on investigation of infective trane- mienione in » unique strain of EZ. coli, yvherein A, Boivin has reported the transformation of « rough variant of one type (C?-R) te the emooth form of a type (Cl-S) from which the active extract is prepared. 1, See McCarty, M., Taylor, H., Avery, 0. 7, Biochemical studies of environ mental factors essential in transformation of pneumococcal types. Cold Spr. Hard, Symp, Quant. Biol. 113 177-182 (1046). ~afle Subtitle #1. Genetic centrol of bacterial enzymes Although the general theory that the primary function of genes is the elaboration of ensymes is now generally accepted, there has been little explor- ation of the genetic control of specific enzyme systems (1), What work has been done has led to the generalization that there is a one-to-one correspondence between genes and enrymes, from which it haa been inferred (2) that the speci- ficity of an enzyme ia conferred upon an inactive precursor in a single step by a single gene, Bacteria have long been favorable materials for enzymatic research; now that genetic procedures are also feasible in E, coli, thie organism becomes vell- euited for the experimental study of gene-enzyme relationships, Carbohydrate splitting enzymes were selected for this study beccuse of the ease with which mutants lacking particular enzymes can be detected, using indicator media, Abdout 200 mutants deficient in the splitting of lactose have been isolated so far, Genetic and chemical tests are etill under way, but it can be stated that mutations at no less than eight different genetic loci can lead to the lowe of lactose-eplitting activity, To a limited extent so far, mutations at aifferent loci can be dintinguished by their effect on the ensymatic splitting of synthetic analogues ef lactose such as mothy]-B-D-galactoside. Tikewlse, at least four genetic loci involved in maltose-splitting enzywes have been identified, "Progressive® mutatione, in the direction of increased enzymatic capaci- ties, have been looked for also, Thus far, in spite of extensive attempts, only two have been found, one e-pable of utilizing the glucoside salicin, and one capable of utilising the straight-chain component of starch, amylose. In view of the work previously described, pert of the difficulty in adapting KF. coli to new substrates may lie in the possibility that several gene changes are required concurrently, -3- Preliminary chemical studies suggest that only a single ensyme is involved in the splitting of lactose to hexose sugars, but it is premature to deny the universal validity of the 1:1 theory, However, to impute eight, and possibly more, enzymatic steps to the reaction is a strain on the imagination. Some modification of the one-to-one generslization may be anticipated, Clarification of the relationships will require an extensive program of chemical and genetic analysis, 1, Beadle, G. ¥, Biochemical Genetics. Chem, Rev, 37: 15-96. (1945) @, Bonner, D. M, Biochemical Mutations in Neurospora. Cold Spr. Hard. Symp. Quant, Biol, 12: 14-24, (19h6) Dr. K. P. Link of the Department of Bicchemistry is kindly asaisting in the preparetion of some of the synthetic substrates to be used in this study, ~-4. Subtitle #2, Genetic transformations in BE, coli Dr, Boivin has Kindly made available etraine Cl and C2 mentioned in the preface, Since the C2R ~ C15 transformation is difficalt to avply quantitatively, and provides only a single marker, we have endeavored first to obtain a series of mutants of both these strains, By the use of etraine carrying several trans- formable characters, such problems as the linkage or association of distinct transforming substances, and the possibility of separating the chemical of spe- cific traneforming agents can be attacked, Several mutants have been obtained in the transformable type C2, mostly nutritional mutants, requiring various amino acids for growth, tncluding arginine, leucine, lysine, threonine, tryptophane, isoleucine, and valine, One strain has been isolated which spoears to require a hitherto unidentified substance, Tests for the induced reversion to normal of these sutants under the influsnee of extracts of Cl are under way as of thie date. 4 G2 mutant which was ansaerogenic (unable to produce OO gas in the course of sugar fermentation) was however shown to be converted to the wild, aerogenic type by such extracts. To date this has been our first confirmation of Roivin's conclusions, Mutants have aleo been obtained in C1, and will be used to determine whether watant characters can also be transferred in this type of hereditary transmiesion. The geneticiet's primary interest in these phenomena reaides in the gene-like behavior of transforming extracts, Not the least of our efforts must therefore be in the direction of purification, separation and chesaical characterization of these agents, This work wae initiated in collaboration with Prof, B. L, Tatum at Yale University, with whom day-to-day exchange of information and atrains ic continuing.