Worm Breeder's Gazette 10(1): 33
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
I am attempting to clone the fem-1 gene, which is essential for male development and hermaphrodite spermatogenesis in C. elegans (Doniach and Hodgkin. Dev. Biol. 106:223. 1984). In the absence of mutator- induced alleles of fem-1, I initiated a walk from a Bergerac (N62) /Bristol Tc1 polymorphism, eP14, which had been cloned by J. Hodgkin, and which maps 0.1 mu to the left of the gene. A region of 60 kb flanking the polymorphism proved to be unrepresented in the cosmid libraries of A. Coulson and J. Sulston, and this region was traversed with clones derived from two of their lambda libraries. The eP14 contig now spans 217 kb, including two cosmid contigs of 70 kb each, and both ends again lie in regions which appear to be absent from the cosmid libraries. The unc-5 gene maps between eP14 and fem-1, 0.03 mu from the latter ( Doniach and Hodgkin, op.cit.), and may provide a more favorable point at which to resume the walk to fem-1. D. Moerman and R. Hoskins kindly provided strains carrying the unc-5 alleles st1000, st1002, and st1003, which arose in the mut-6 background of RW7096. DNA isolated from each strain after 6-10 outcrosses to N2 contained 2-5 Tc1- containing EcoRI fragments not present in N2. Two of the novel Tc1 elements in st1003 cosegregated with unc-5 in 24/24 recombinants in the dpy-13-unc-5 interval and 18/18 recombinants in the unc-5-bli-6 interval. Both elements were absent from a revertant of st1003. One, eP52, has been cloned as a 5 kb EcoRI fragment. Sequences flanking the Tc1, when used to probe Southern blots, detect a different polymorphism between N2 and each of the mut-6-induced and between each allele and a revertant of that allele. This observation leads me to believe that I have cloned part of the unc-5 locus. The st1003/N2 polymorphism can be accounted for by Tc1 insertion at an EcoRI site, resulting in destruction of that site. Simple insertion of Tc1 cannot account for the pattern of hybridization in st1000 or st1002, although the polymorphic fragment in each case includes Tc1. None of the patterns seen in the revertants can be explained by simple excision of Tc1, and none is identical to the wild-type hybridization pattern. More accurate definition of the extent of the gene and more adequate explanation of the rearrangements observed in the mutants and their revertants should be possible when I have cloned DNA spanning the locus. To this end, and to continue the walk to fem-1, I have identified several clones in a lambda library of C. elegans DNA which hybridize to an eP52-derived probe.