Vol. 35, 1949 GENETICS: J. LEDERBERG 183 



between the two Lac loci, bringing the -f- alleles into coupling phase will 

 be lactose positive. That these cultures are segregating for two Lac — 

 factors has been confirmed by physiological and genetic tests on the 

 segregants. 



Heterozygotes from Standard Crosses. — All the heterozygotes so far re- 

 ferred to are the issue of crosses involving the hypothetical "Het" factor 

 derived from H-l. In these crosses, several per cent of the prototrophs 

 are demonstrably heterozygous. Previous and current controls showed 

 that, if they occurred at all in "normal" crosses, they must be very much 

 rarer. In order to make a more thorough test, advantage was taken of 

 the very close linkage (less than 1% recombination) which has been 

 observed between the Laci and Lac4 loci mentioned above. B — M — Laci — 

 Lac 4 + was crossed with T — L— Bi — Laci+Lac 4 — . Much less than 1% 

 of the prototrophs of this cross on EMS — lactose are Lac + . It was reasoned 

 that a Lac-f- prototroph might represent either a very rare crossover, or a 

 diploid in which the + factors were carried on opposite chromosomes. 

 Because most haploid recombinants, being Lac — , can be set aside by 

 inspection, this is a fairly efficient way of screening for rare diploids. In 

 this way, heterozygous diploids were also obtained from normal stocks, 

 not carrying "Het," but they constitute only about 0.1% of the proto- 

 trophs. These prototrophs are generally similar to the previous ones, 

 except that they are somewhat more stable. It has not been established 

 whether new "Het" mutations have occurred here. When these diploids 

 are singly heterozygous for various sugar fermentation factors, they also 

 show non-random segregations, but these mutations are not the same as 

 those in the "Het" series, so that a direct comparison is not yet possible. 



Discussion. — It is not clear what relationship there is between the 

 exceptional persistence of these diploids, and the abnormalities in their 

 segregation. There may be some direct connection between their hetero- 

 ploidy, i.e., deficiency for the Mai region, and the prolongation of the 

 diplophase. But the very disturbing possibility has not been discounted 

 that these heterozygotes merely reveal a situation which also operates in 

 the formation of haploid prototrophs from transient zygotes. If this is 

 so, the linkage map of E. coli K-12 will have to be systematically re- 

 examined, with the use of several unrelated sets of stocks. In any case, 

 the discrepancies affect only the details of chromosome behavior. These 

 heterozygotes, on the other hand, have provided an unexpected con- 

 firmation of the sexual basis of genetic recombination in this bacterium. 



Although the heteroploidy is somewhat of a limitation, heterozygote 

 formation is a very helpful tool in genetic analysis. Since all types of 

 recombinants, not only prototrophs, can be recovered, it is feasible to 

 extend pedigrees to several generations, and interesting combinations of 

 factors can be put together in a form allowing their use in crosses. For 



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