Worm Breeder's Gazette 11(2): 79
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 continuing characterization of unc-22 mutations caused by trimethyl-psoralen (TMP) as part of an effort to determine whether this mutagen produces a different mutational spectrum than EMS. TMP, activated by near-UV light, cross-links DNA, so we might expect a high frequency of rearrangements and deletions as a consequence of breakage and repair. Preliminary analysis of 22 homozygous-viable unc-22 mutations by Southern blot gave the following distribution: Deletions: 8 Possible Inserts: 1 No Detectable Polymorphism: 12 The probes used were DM18 and DM20, which cover most of the transcribed region and 10kb of 5' upstream sequence. (DNA probes kindly provided by Carol Trent.) Sizes of the 5 internal deletions ranged from 100 bp to 3.5 kb; the other three represent a 5' and two 3' end fragment deletions. One of these (5') appears >10 kb. We plan to extend this analysis using cosmid probes to determine whether any of the mutations showing no polymorphism represent deletions of sequences not covered by the probes used so far. Mutagenesis of L4's ( 60' light at 300 w/cm2 following a 15 min TMP soak) gave a mutation rate of approximately 10+E-4, comparable to EMS rates for unc-22. In addition to looking for homozygous mutants, we screened F2 plates for heterozygous twitchers representing homozygous inviable mutations, suspecting that large deletions in the unc-22 region might include nearby essential genes. We retrieved 9 such mutants in a run producing 15 homozygous viable mutants. Several of the former are currently being balanced for analysis. None extend to dpy-20. Thus TMP appears quite efficient at generating small deficiencies, which may make it a useful mutagen for producing null alleles and for molecular tagging. Protocol available; we would be interested to know if it works on your favorite gene. [See Figure 1]