Worm Breeder's Gazette 11(2): 61
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.
To facilitate the molecular cloning of bli-4(I), we have screened for and obtained a spontaneous allele in a mutator strain. Because the most frequent phenotype of bli-4 alleles is early larval arrest, we assumed that a mutator allele would be lethal (ten of the bli-4 alleles are lethal compared to one visible, e937, which is a blistered adult). A mutator strain of the genotype unc-63(e384) 50)(I);mut-6(IV) was constructed and designated KR1822 (mut-6 is from RW7097, obtained from D. Moerman and R. Waterston). KR1822 segregates spontaneous twitchers at a rate of 3x10+E-4. Heterozygous males of the genotype dpy-5 7) ++ were mated to KR1822 hermaphrodites and the F1 progeny screened for blistered worms, presumably of the genotype unc-63+ )dpy-5 7) +. These worms are blistered because bli-4( lethal)/bli-4(visible) worms are severely blistered. Three Bli worms were identified after screening 82,300 chromosomes: one male that did not out-cross; one hermaphrodite that died without progeny; and one hermaphrodite that survived. The frequency of bli-4 induction was 4x10+E-5, or about ten-fold lower than twitcher induction. The mutator allele has the following characteristics: 1) it is lethal, as expected; 2) it fails to complement the other bli-4 alleles; and 3) it is linked to unc-63 and unc-13. The allele has been designated h1010 and was balanced by the translocation sZT1 in the strain KR1858. To identify the insertion site of the transposable element we probed KR1858 DNA with 10 cosmids spanning an interval from hP5 to the left breakpoint of hDf8. hP5 is 0.3 mu to the left of bli-4; the hDf8 breakpoint is to the right of bli-4. We chose to use EcoRI and SalI to digest the genomic DNA because EcoRI does not cut inside Tc1, and would produce a novel band 1.6 kb larger than in the parental strain. SalI does cut within Tc1, and would produce two novel bands smaller than that of the parental strain. Only one of the cosmids, K04F10, detected band shifts: a 1.4 kb to 3.0 kb band shift in EcoRI digested KR1858 DNA; and a 21 kb to 16 kb and 6 kb band shift in SalI digested DNA. This pattern is consistent with the pattern of band shifts predicted for Tc1 insertion. We are now in the process of confirming that K04F10 includes bli-4 by microinjection-rescue.