Worm Breeder's Gazette 12(1): 22 (September 1, 1991)
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 trying to clone and sequence the muscle gene unc-60 ,located on the left end of LGV. The genetic organization of the unc-60 region has been described previously by Kim McKim (McKim et al., 1988). Subsequently, cosmid microinjections were performed and a rescue was achieved, localizing this gene to a deleted 22 kb cosmid, F53E2 (M. Wakarchuk, WBG Vol. 11, #1).
To identify the unc-60 sequences we have tried two approaches. First, genomic DNA prepared from N2 and three gamma induced alleles was digested with HindIII, SalI and EcoRI and blotted. These Southerns when probed with the entire cosmid were uninformative.
The second approach was to identify sequences conserved in C. briggsae. We have constructed a restriction map of the cosmid. Southern blot analysis using various cosmid subclones has shown that there are several classes of repetitive DNA within this cosmid. Subclones containing these repetitive sequences hybridize to many C. briggsae fragments at low stringency. A 5 kb PstI fragment was found that contains no repetitive elements but exhibits C. briggsae homology. This fragment was used to screen 30,000 phage plaques from a lambda ZAP cDNA library (a gift from R. Barstead and R. Waterston) resulting in the isolation of 12 cDNA's which range in size from 500 bp to 1500 bp.
The 5 kb PstI fragment and an overlapping non-repetitive 6 kb HindIII fragment are now being used for microinjection in attempts to rescue unc-60 (M. Wakarchuk per. com.). As we are unsure of the number of coding elements within the cosmid additional cDNA screens are currently underway. Sequencing of the repetitive DNA and the cDNA's will follow.