ew) Separate mention of the cultures that were classified as double (-) by transduction test mst be made partially because the results are more incomplete and partially because they may offer some additional information upon the transduction phenomenon. Four such (—) have been obtained, three of the gal)-gal2- type and one of the galo-galy- typee- The evidence that such cultures are (--) is that they are transinced either by homotypic xor hetero- typic lysates but are transduced by wild type or some other gal (-). Lysates of these (--) cultures have been found to have little transducing activity regardless of the gal (-) tester used with but one excep ti oh. Whether this implies a failure of the phage particles to pick up a fragment of cell chromosome or whether the resultant thansduction is not phenotypically (+) through some interaction among the genes Concerned is not known. The exceptional case resulted in the recovery of each of the (-) making up the (—-) wauxrecsysrsd individually and not conjunctively. The homotypic locks transduced with this lysate was not recovered among the segreganta. As might be expected the (—) are more stable on galactose medium and have seldom been seen to revert. 2 Some experiments of interest have been performed with one of the (—) obtained. (It wa ortunately’a prototroph and the results obtained with must é it mtkx also be repeated and extended with auxotrphic strains. Although this (—-) was not transduced by uttkex , lysates of wither (-) singly it wae transduced to a lesser extent ( where a solid layer of papillae by a ratture of tue 190 with a (=) weuld have been obtained, less than 100 papillae were f ound),. In this ae : wos 4 case it, taken that the celle transduced to (+) had received two phage particles se meats with the addition of two (+) alleles in separate pieces. The cell that was transduced ty (+) may be represented as follows: 2 le and the resultant transduction as follows: a a ~27—-1t. -2t.~] ~~ In this case the extra (~) added in the segments are inferred from the results with transductions of single (-) in which the heterotypic lecus is recovered among the segregants. aus Segregation from this transduction in the absfnce of crossing over or exchange between chromosome and segments can result in three types of (-) segregante, | (1) meat ae (2) eee (3) 2-1 &2---1*. -2t-~1-- which would be classified as (—), (27) and (1~) presumably. With exchange between segments and the chromeome segregants with the (+) allebes weuld be found in the chromsome and subsequent segregation would yield( in addition to the types 2 and 3 above with the (+) transposed) the following types: (tb) e271 an (§) —~-2t-—-1"———s An additional type can be obtained if there be exchanges betteen segments. The order of frequency of exchange and segregation of the ebove types is unknown but on anaslogy with the simple tranifupions the first thewe mentioned would be expected most frequently, that is, loss of » segment is more frequent than exchange and loss of a segment.(This in turn is dependent upon the independence of exchange and loss) Examination of 2 separate segregants from one such transduction gave the following distribution of segregants by transduction test: 13 (—), 6 (17) and 5 §27). Since over 50 percent of the segregants were (—) it appears that when loss of a segment occurrs it is more likely to involve loss of both segments. The (17) and (27) found could be of two types, 2,4 and 3,5 above respectively. These types can be distinguished by means C of analysis of (+) reversions. In cases 2 and 3 the reversions will be unstable and segregate, and in cases 4+ and 5 they will be stable for galactose. Reversions were examined for their stability from each of the (-) obtained. All the (17) WEEE gave stable reversions and therefore were presumably of the ---27--1"—« type. Cf the (27) examined all but one gave stable reversions and therfore the two typee——-2°——-1?—» and 2 * were indicated with the most frequent being the former, Examinatioh of the ts (2) culture giving the unstable reversions showed that it xpaht did segregate (—-) cells but as yet it has not been established that it segregates (27) of the following type -——-2-—l*—* , The reversions of tty the type 2 (27) can be of two types and they should (perhaps) be distinguishable in turn by the segregsats that ty yield. Reversion of the form ~--2 yoy ---* should be expected to segregate (—) predominately ‘and veverstee ct ae the form —--2 ho shnvuld be expected to segregate (1) predominately. ea Reversions of the type 2 (27) appear to be of two types. From one type 33 segregants were obtained, of which 32 were (--), the remaining one 6 (27). The other type gave almost equivalent amounts of (2) and (—) and no (1°) thus far. The failure to recover (17) types from the wrx reverted cultures is disturbing but this may be related to elimination of the gal, locus in crossés. Presumably crosses between naan aa and —--2t—]"-—-« should yield a larger number of (+) than crosses between (17) and (27) of normal constitution when there is sucessful transfer of the segment through the zygote. these (+) in addition would be unstable for galactose. The culture used unfortunately is a prototroph and unless sucessful crosses between it and a Hfr strain can be accomplished the problem can not be attack from this aspect. ( Sucessful transmission of the eegnent thr ough the zygote was observed in some early experiments not related to the adore, 6G Examination of another (—~) has begun. In this case Gal, and aly, are involved and a croseable stock has been selected. There has been another complication in this case. That 4s when the culture was first isolated,and also in the case of a repaat test, it was not found to be transduced by either (27) or (4°) lyssates. Infeveral additional tests it has also reactive in this manner. In the instances where it was attempted to obtain transductions by mixtures of the two lysates it was found that the culture was transduced, to a lesser extant, by lysates of (27). Temctupxtustuxesrexcactrmmeserartiz It was thought to explain this incongruent result by postulating that reversions had occurred turing the growth of the culture and that in effect the cultge consisted of (—) abe} (47) contaminants. On this assumption the gransduetions of the culture would in effect be of the form (2-) ——-x (47) and the resultant transductions would be expected to segregate (4”) predominately. This was not the case, of the six segregants examined(from six separate transductions) 3 were (27), 2 were (0 and only one was (4"). This does not rule the explanation om but requires a frequency great eat of exchange between segzuent and chromosome for compatibility. \ Yxamination of this culture had progressed to the stage of isolating. a (4°) segregaat that gave unstable reversions as well as a yuxtet type which did not, at the time of writing. Not all of the Gal- cultures studied have been found transducible alz though the most frequently occuring (-) after ultraviolet radiation appear to be of this type. Three difinctly differ sat occurrence, of non-transduci ble gal- have been found. Two of these were induced by ultraviolet, and the third by copper exposure ( H. Buyers). ‘ne of the ultraviolet mutabis has been examined to some extent. The results are given in table 18. It appeare that this (-) is not transduced by any of the lysates and futher that lysates of it in turn traifyfuce all known transducible loci, but Galo with lowered frequency. QS) Table 18 Analysis of a Not-traneducible Galactose Locus in W2312 hy Tranaductiqn Asaay Experiment Plate Additions None : HFT Lysates AFT Gal. - Gal,~ Gal)- Wild Type 206 (1) o* o* o* 0 - (2) 0 8 - 7 9 220 (1) 0 0 0 0 - (2) 0 . o** o** ore 0 * mmber of papillae per plate ** NET (normal frequency of tranaduation) lysates used in these cases y fable 14 Activity of lysates of W2312 ou Selected Galactose Loct Galactose Plate Addition Locus | None W2312 Lysate Gal,- Lp* ips . 378 a7 ip” (220) 8 ? (221) 19 28** Galy- Lp* 17 74 Galg- Lp® 3 - 121 " mumbers of papillae per plate #9 12/12 examined were found to be stable Galt fable 15 Results of Crosses of W2312 with Selected Galactose Loci Selected Galactose Numbers Locus Gal+ Total Prot otrophic Rechmbinants Percent Gal+ Galo~ x 1 2112 0.05 Galy- F* 1 196 0.5 ® For the purpose of collecting new gal- and for observing the occurrence of tumnsducible loci two separate experiments were performed, Gal- mutations were induced in ¥1673 (glyc or ser)~ prol” and W1765 hist™ leuc” by means of ultravinlet. Table 19 gives a summary of these experiments. Reccurrences of both Gal,- and Galo- were found as well as a number of new loci and pessibdly several (-—-). Ne recurrences of Gal,- were observed. The effect of ultraviolet radiation on the transducing activity of lysates has been investigated in three experiments. The firet two experiments were concerned with MFT lysates, the last with an HFT lysete. The effect of ultraviolet upon NF lysates is shown in figure 2. With increasBing dose of ultraviolet there is a linear increase in the activity of the lysates on Lp* or Lp* assay cells until a survivial of the plaque-forming titker has become reduced about 1073. Thereafter there is a gradual decease in transduction activity with increastog dose. On Lpe there is a slight increase in transducing activity ant then a gradual decrease. The maximum reached by the lysate™ on Lp* or Lp celle is about four times the maximum reached on lp® celle. In performing this experiment about 16° Ip® assay celle were used, since figure 1 indicates that this number of cells may {nitcate only about x one-thizd to one-fourth the number of trensductions actually present the Lp® assay is probably thet much low. fhis then would suggest that the absolute number of transductions 1s approximated uponp® cells when a sufficient mumber of cells are used and that the action of ultraviolet is to increese the assay pn Lp* or Lp* cells to the level of the absolute number present. In connection with this it should be noted that survival of the transducti onexsuxy Lp® is still about 0.5 even at the extreme doses used. From the above it 4s suggested thet the action of ixsxtes of ultraviolet is weveral fold. Fires and most rapid fs the destruction of plaque forming activity a Lp* cells. Secoxmly, to destroy that property of the ag receds Bede che phage which causes them to pe"excluded" by lysogenic celle, and thirdly to destroy & oy, i - Aah Oe, Table Transduction Assay of Some Galactose Negative Mutana&s Induced by. Means Ultraviolet . Possible Culture Mutant Transduced by HFT PESHEMIE Treated Designation Gail,- GRTo— Gal)- Genotype W1673 Lp® W231o 0 + 0 Gal, -Galy- W2311 0 + 0 " " W2322 0 oO 0 nontransducible W2313 + 0 + Gal>~ Ww2314 + + + Gal - W2315 + + + Gal_- W2316 0 + + Gal, - W2317 0 + 0 Gal, -Gal,~ W2318 0 0 0 nontransducible W1765 Lp® 238-2 0 0 0 nontransductble pang + + + Gal _- 238-6 0 + + Gal,- 238-8 + + + Gal_- 238-10 + + + Gal,- 238-11 0 + 0 Gal, -Gal,,- 238-12 + 0 + Gal, 238-13 + 0 + Gal 27 G the transducing activity itself, perhaps by destroying the adsorption of the phage particles. The effect of ultraviolet on HFT lysates is similar to that of UV on KFT lysates. The increase in transducing activity with dose in this case ig not as great as with NPT lysates. A maximum is reached that is approximately equivalent to the plaque titer of the lysate which suggests that plaque and transducing particles may be the same but that appearance of a particle as a plaque exchudes its appearance as a transduction. Platings for plaque formation on EMB galactose have not indicated that one particle can function in both capacities but the appearance of a plaque might be obscured by papillae formation. The sum of the activities (maximal) of the lysate on the two assay loci is 2-3 times the plaque sktkxmx titer, which may be an indication that the activities are confined to a single particle. The occurrence of transductions with Lyp™ genotype has been noted with this lysate, and the equivalence of plaque and transduction titer might not be expected on the aswumption that in these cases the effect was accomplished by a defective phage particle which would not give as well as to rise to plaques 3F lysogenization. (This would reqbhire that Lp’ genotypes were the result of such defective particles rather than of a defective act of lysogen- 4zation.) SOON O t 9 “oogrh OM F ag ‘O09 N|3OZ131I0 3N3ONF ” q ayes lt ‘S¢n NI JOWW “OOoOrR OM YY SCHRTELT a7 9OR O H v “ N Oo Suu p4gonps tea e4 HON! 43d SNOISIAIO OF K S3154A0 GS DIWNHLIaVvVOOT-IWSS UBdvd HAVYD NSOZL3IA OCISI-OPE "ON (< v “gor on 2 ca ” 10 Ultaviolet Dose in Hiuates s Ss — nt TOONnOM ¢ g 7qns masp mip earl “fhf AAs APQUIAN } ‘O90 NAOZLIIG 3N39N3 , HON! Yad SNOISIAIO Zi KX S3IDAD G JINHLIaV904-INSS N3dVvd HdV4eH NADZLIIA SIS1-Orvre ‘ON gre pla ' S) Gace ad gerves ted as 2 uv. 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