Worm Breeder's Gazette 12(2): 37 (January 1, 1992)

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.

RFLP (Restriction Fragment Length Polymorphism) and Repeated Sequences of Caenorhabditis elegans

Hiroaki Kagawa, Hideki Ando

Department of Biology, Faculty of Science, Okayama University, Okayama, 700 Japan

We are cloning kra-1 (ketamineresponse abnormal) gene together with unc-68 and mec-1 genes on LG V of Caenorhabditis elegans. One of a Tc1 fragment from the mec-1 mutant; TU952 was located at the reasonable position on the physical map (WBG,#11,No.2). On the process of screening to find any RFLP bands in isolated mutants, a unique band appeared in unc-68 ( e540 )on a Southern analysis by using cosmid clone W02D7 as a probe. Four strong bands; 11kb, 5.7kb, 3.6kb and 2.6kb (sizes were tentative) were also stained with this probe. The size of a unique band of unc-68 ( e540 )mutation was about 3kb shorter than the corresponding band of N2 .Extensive Southern analysis showed that a unique band also appeared in other unc-68 mutation( x14 and X24 ).This means that a unique band might not correspond to the mutation.

Surprisingly this band was also found in N2 worm which is keeping in the Hosono's lab (Kanazawa, Japan). N2 of our lab comes from MRC. It is not clear why two N2 worms(MRC and through U.S.) had the different band. Some person should solve the reason.

The band pattern of strains with gamma-ray-induced alleles of mec-1 ; TU1022 , TU1029 , TU1041 , TU1044 and TU1046 ,were the MRC type. Strains with TR679 -derivedmutations; TU295 , TU952 , TU941 and TU692 had RFLP bands which might come from the reason of outcross. The second band (5.7kb) was the same size which located exactly in the cosmid W02D7 .Other three bands were not found in the cosmid clones of this region. To confirm which fragment hybridized to these four bands, restriction fragments were analyzed. Finally the repeated sequence was found in the 2.6 kb fragment of KpnI digest of W02D7 and had the nucleotide sequence of conserved 14bp AAGGTAGTCAATGA and variable 3-4 bp (T/G)C(A/T)(A/G/-). This 17-18bp repeats were tandem and more than twenty. Northern blot analysis is also progress. Cosmid W02D7 encoded three transcripts. It could be possible to locate these repetitive sequence on the chromosome. Preliminary screening of a YAC filter showed that more than twenty places were hybridized to KpnI-2.6 kb. There might be another repeated sequence in this fragment. Although we can't completely analyze this fragment, this repeat sequence can be used as a probe for study of RFLP and may effect gene expression or gene duplication.

Correction: The tropomyosin gene, tmy-1 of Caenorhabditis elegans locates on LGI just left of the unc-54 gene, not on LGV (the last issue of the worm gazette, Vol.12, No.l)