Worm Breeder's Gazette 14(4): 46 (October 1, 1996)
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.
Department of Biochemistry and Molecular Biology, University of New Hampshire, Durham, NH 03824
The oncogene src encodes a tyrosine kinase implicated in signal transduction pathways that control cell growth and development. When activated by mutation, src can induce neoplastic transformation. Despite intensive study for many years in a variety of organisms (almost exclusively vertebrates), a good understanding of src function still eludes us. We are trying to knock out src-1, a C. elegans homologue of vertebrate c-src, in order to characterize the role of src-1 in C. elegans development. Such knowledge will contribute to the understanding of src function in all organisms. We have isolated a src-1::Tc5 mutant using the PCR / sib-selection screen of Rushforth and Anderson (Zhang et al, C. elegans Meetings Abstracts - 1995). In this mutant, Tc5 resides within the fifth intron of src-1. The presence of Tc5 does not confer a visible phenotype because it is spliced out with the intron leading to normal mRNA production. Efforts to isolate a deletion caused by Tc5 excision are in progress. src-1 resides on a small "orphan" contig that has not been placed on the physical map. The mutant we isolated provides the opportunity to map the gene via a PCR based approach using the src-1::Tc5 allele as a molecular marker. Using a primer within Tc5 and a primer in exon 6, we are able to specifically amplify a 1.2 kb product only from individuals heterozygous or homozygous for src-1::Tc5. We crossed strain TW399 [src-1 (cj290::Tc5)] by strains containing markers near the center of each linkage group: LGI dpy-5, LGII dpy-10, LGIII unc-36, LGIV unc-22, and LGV dpy-11. Individual F2 animals homozygous for the marker allele were picked and screened by PCR for src-1::Tc5. In each case, approximately 75% were positive indicating that src-1 was not linked. We established that src-1 is not X-linked by following its inheritance through two generations of XO males. We continued our mapping efforts by crossing strain TW399 by strains containing markers located on either arm of each linkage group: I L bli-3, I R unc-54, II L sqt-2, II R unc-52, III L dpy-1, III R bli-5, IV L dpy-9, IV R dpy-4, V L unc-34, and V R unc-51. In each case but one, approximately 75% of individual F2 animals homozygous for the marker allele were positive for src-1::Tc5. The one exception was F2 animals homozygous for bli-3, a marker on the left arm of linkage group one. Only 5% (3/60) were positive for src-1::Tc5. Control reactions with the same DNA confirmed that the negative reactions were in fact due to the absence of src-1::Tc5. Hence we conclude that src-1 is located near bli-3 on the left arm of linkage group I.