Worm Breeder's Gazette 11(4): 14
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.
We are in the process of cloning the age-1 gene. The most extreme mutant allele in this locus has an increase in mean life span of 60% and in maximum life span of about 110% at 25 C. In backcrosses to N2, the age-1 locus was tightly linked (<1 map unit) to fer-15 and cosegregated with a four-fold reduction in hermaphrodite fertility ( Brd), as well (Genetics, 118:75, 1988). fer-15 and emb-27 have been mapped to the same interval in the subcluster on chromosome II by deficiency mapping, which we have confirmed. We decided to employ a strategy for cloning age-1 wherein we would identify RFLPs generated by the endpoints of specific deficiency chromosomes and identify fer- 15, d injections. However, recent data suggest that age-1 is separable from fer-15 and nearer to unc-4 (see Hutchinson and Johnson, elsewhere in this Newsletter). Nevertheless, we have decided to finish up our molecular analysis of this region of chromosome II. Three deficiencies have endpoints immediately adjacent to the left of the fer-15, hile eight deficiency endpoints mapped adjacent to the right. We have identified all three endpoints to the right and seven of the eight endpoints to the left. Along the way, we've also identified the other ends of four of these deficiencies and identified a possible deficiency associated with fer- 15(b26) in T13F9. This information is summarized in the figure. We chose cosmids in the Rol-6 contig, which covers most of chromosome II, based on Phil Carter's observation that C26D10 revealed an RFLP in mnDf104 (WBG, 10,2:60) which we confirmed. Since the deficiency chromosomes must be carried in the heterozygous state, we anticipated that an endpoint would be most reliably detected as a new band in a genomic blot. (This is because a band that has a two-fold decrease in intensity is hard to verify without densitometry and likely to be variable for different deficiencies.) Most of these endpoints have been verified as giving new bands with more than one enzyme (to distinguish deletions from single-base changes). In the figure below, the relative position of these endpoints within a cosmid are unordered (we don't yet have a reliable restriction map of the region). We have not yet probed all combinations of deficiencies and restriction enzymes with all overlapping cosmids so we can't localize endpoints to cosmid subregions. As a control for rearrangements, we have been routinely comparing band size of genomic blots with band size of cosmid digests and generally find good correspondence. A few cosmids (e.g F58D3) were difficult to recover intact, often giving identical deletions in different picks. Our current plan is to prove that the fertility deficit is separable from age-1 and to confirm our identification of fer-15 and emb-27 by using T13F9 and adjacent cosmids to rescue these mutants by injection. We hope to get back to cloning of the age-1 gene soon and leave fer-15 and emb-27 to others ( Steve L'Hernault has similar, but not identical data). [See Figure 1]