Worm Breeder's Gazette 10(1): 32
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.
Mutations in daf-1 convey a dauer-constitutive phenotype with a maternal effect. We found two apparent Tc1 insertion alleles of daf-1 using the mut-4; background derived in the Waterston laboratory. One spontaneous revertant was also selected for each of the two alleles and the sites of reversion were shown to be closely linked to daf-1 by appropriate genetic crosses. Also, the revertants did not exhibit a dauer-defective phenotype, indicating that they did not harbor new dauer-defective mutations epistatic to daf-1. To facilitate detection of novel, daf-1-associated Tc1-containing restriction fragments, strains with lower Tc1 copy number were derived by crossing to Bristol backgrounds. Genomic DNA from lower copy number strains was probed with Tc1, revealing a 2.5 Kb BamHI fragment present in both mutants and absent in the one revertant examined. This fragment was recovered from an agarose gel and cloned in pUC18. Subcloned flanking sequences were used to probe genomic Southern blots of the non-outcrossed mutant, revertant and parental mutator strains. The flanking sequences hybridized to a 2.5 Kb BamHI fragment in DNA from both mutants, corresponding to the fragment that was cloned and to a 0.9 Kb BamHI fragment in the parental mutator strain and in the examined revertant ( corresponding to absence of Tc1). The hybridizations also showed that the flanking DNA is single-copy. DNA sequencing has revealed an open reading frame on both sides of the Tc1 insertion and a putative intron of ~50 bp. An initial search of the sequence database, however, has not revealed any interesting similarities with known genes. Analysis of the complete wild-type gene may produce further information. Maybe it will be something new and different.