Worm Breeder's Gazette 14(5): 32 (February 1, 1997)
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.
Albert Einstein College of Medicine, Department of Molecular Genetics Ullmann 703, 1300 Morris Park Ave. Bronx NY 10461
In the process of genetic mapping we learned that the left arm of LG IV contains restriction fragment length polymorphisms between the strains N2 and DP13 (a Bergerac RW7000 subclone). These were detectable by Southern hybridization with cosmid probes (G. Beitel, personal communication). This region happens to be the target of current sequencing efforts. Visual inspection of cosmid sequences revealed the frequent occurrence of repetitive DNA, leading us to wonder whether the RFLPs arise from differences in lengths of these repetitive sequences in the two strains. The repetitive sequences are similar to satellite repeats in that they consist of tandem repeats of short units flanked by nonrepetitive sequence. We analyzed 12 such repetitive regions, with repeat unit lengths varying between 6 and 30 bp, and between 300 and 1800 bp in total length. PCR was performed on single N2 and DP13 worms as described by Williams (Epstein & Shakes, Methods in Cell Biology volume 48) using primers based in nonrepetitive flanking DNA. Analysis of PCR products by 1% agarose gel electrophoresis showed length differences between N2 and DP13 for 7 of the 12 regions. In each case the length of the fragment amplified from N2 was consistent with sequence data. DP13 PCR products were longer or shorter than the corresponding N2 PCR products by a few hundred base pairs. It seems likely that the length differences are due to differences in the number of unit repetitions. We plan to test other strains for these polymorphisms. PCR analysis of polymorphisms is a simple and quick way to determine the genotype of a single worm at a physically defined location. This method can be useful for gene mapping through analysis of F2 progeny from a cross between a worm homozygous for the gene of interest and a wild-type worm of a different strain (Korswagen et al., PNAS 93, 14680-14685, 1996). The distance between a repeat region and the gene of interest should be inversely proportional to the frequency of coincidence of the mutant phenotype and homozygosity for the repeat length of the strain of the mutant parent.