Worm Breeder's Gazette 10(3): 71
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.
The set of cosmids that constitute most of the physical map of the worm at present cover an estimated 90% or more of the genome. Unfortunately the residual 10% may include some important genetic loci. One such appears to be the region including the major sex determining gene tra-1, on the right arm of LGIII. As described in the previous issue of the Gazette, a 200 kb YAC (yeast artificial chromosome) Y21B3 hybridizing to the cloned Tc1 insertion eP1 also detected a small contig, which was found to be located in or adjacent to tra-1 itself. The contig was not oriented relative to the genetic map, but several natural polymorphisms from the Freiburg wild strain RC301 were found on the contig and used to provide an orientation. Phage, cosmid, and YAC libraries were probed with the leftmost fragments of this contig. Small extensions were obtained by phage walking, which were used to identify additional rearrangements in several tra-1 loss-of-function alleles, for example a 13 kb deletion in the strong (class A1) allele e1834. An additional extension was obtained by probing a phage (lambda 2001) library plated on CES200, which is recB- recC- sbcB-. The phage recovered did not plate on the standard rec+ host used here, K12.803. Four additional large YAC's were also obtained, from the new library constructed by Waterston et al. One of these, Y48A6 (300kb), was shown to extend only about 30 kb into the existing tra-1 contig, and therefore should extend 250kb leftward, probably covering the rest of tra-1 and extending as far as pha-1. Mutations of pha-1 (formerly sot- 1, 'sore-throat') obtained by Heinke & Ralf Schnabel lead to a failure of pharyngeal differentiation during late embryogenesis (1987 Meeting Abstracts, p.46). The X-ray induced mutation e1855 fails to complement both pha-1 and tra-1, and behaves as a deficiency with one breakpoint right of tra-1 and one breakpoint in pha-1 (because recombinants have been obtained between e1855 and other pha-1 alleles). Y48A6 also hybridized to two new small contigs. Cosmids from these, and from the tra-1 contig, were used to probe wild-type and e1855 DNA. These blots showed first that e1855 is indeed a deficiency (rather than an inversion), probably for the whole tra-1 region, because the right break point has not been found. Second, one of the new contigs is outside this deficiency (eDf20), presumably to the left, and the other new contig appears to span the left breakpoint of eDf20. Y48A6 was labelled and used to probe a genomic library plated on CES200. Of 115 phage recovered, about 20% plate poorly, or not at all, on K12.803. Some, but not all, of these 2094 plate well on the recD host DB1316. The phage have not yet been fully analyzed; it will be interesting to see if they cover the whole of the pha-1-tra-1 region, or whether there are still gaps. Notice that of approximately 500kb corresponding to the sum of Y48A6 and Y21B23, only about 150kb are covered by cosmid contigs. [See Figure 1]