RECOMBINATION ANALYSIS IN MICROBLVL SYSTEMS 67 



binants will be markedly reduced. It is the points of attack nearest the 

 mutated site of the recipient cell (positions 2 and 3 in Figure 9) that 

 give rise to most wild-type recombinants when the recipient cell's site 

 is a point mutation; most of these recombinations will be mutant, not 

 wild, when the site is extended, because of the large number of returns 

 within the mutant sequence. In Figure 9, the only effective recombina- 

 tions will be those beginning in positions 2 and 3 and ending in posi- 

 tions 6, 7, and 8. 



The same depression in frequency of wild-type recombinants is 

 observed irrespective of whether the mutant sequence in the recipient 

 cell is to the right or to the left of the mutant site of the donor. This is 

 why the assumption of polarity is unessential. Thus the lack of agree- 

 ment between recombination frequency in reciprocal crosses is per- 

 fectly understandable in terms of this model. 



There are, however, conditions under which extended mutant 

 sequences will give identical results in reciprocal crosses. First, if both 

 the donor and the recipient cells are marked with extended mutant 

 sequences of approximately equal size, not only will reciprocal crosses 

 give equal frequencies of wild-type recombination, but also these fre- 

 quencies will be extremely low. Consequently, the fact that mutants rrij 

 and m,i show no recombination in Table I does not necessarily mean 

 that they are overlapping. Second, reciprocal crosses between a long 

 marker and a point mutation will give indistinguishable numbers of 

 wild-type recombinants if the most frequent length of sequence in- 

 volved in recombination is very large with respect to the length of the 

 extended marker. However, the very fact that the size of the mutated 

 site is clearly influencing recombination frequency could mean pre- 

 cisely that the length of donor sequence most frequently inserted in the 

 recombinant structure is small— of the same order of magnitude as the 

 length of a small mutational site. 



If we now examine Table I, we see that all markers give asym- 

 metrical results with mutant m,,. In terms of the model, this means that 

 all markers are longer mutant sequences than m,; and are large with 

 respect to the most frequent length of sequence involved in recombina- 

 tion. We should furthermore be able to order the mutants with respect 

 to size, according to the degree of discrepancy observed in reciprocal 

 crosses. To do this, we can consider all of the crosses involving any 

 single mutant both as donor and as recipient. We can then calculate for 

 any given cross of x by y the ratio of the recombination frequencies ob- 

 served when the mutant is the recipient and when it is the donor. For 

 example, for mutant g: 



f(gVPNAo) ^,g^_^^ HgbyDNAM ^^^^^^^^ 

 f(cbyDNAg) {(/ibyDNAg) 



