TARGET-SELECTED GENE INACTIVATION IN CAENORHABDITIS-ELEGANS BY USING A FROZEN TRANSPOSON INSERTION MUTANT BANK

To understand how genotype determines the phenotype of the animal Caen orhabditis elegans, one ideally needs to know the complete sequence of the genome and the contribution of genes to phenotype, which requires an efficient strategy for reverse genetics. We here report that the T c1 transposon induces frequent deletions of flanking DNA, apparently r esulting from Tc1 excision followed by imprecise DNA repair. We use th is to inactivate genes in two steps. (i) We established a frozen libra ry of 5000 nematode lines mutagenized by Tc1 insertion, from which ins ertion mutants of genes of interest can be recovered. Their address wi thin the library is determined by PCR. (ii) Animals are then screened, again by PCR, to detect derivatives in which Tc1 and 1000-2000 base p airs of flanking DNA are deleted, and thus a gene of interest is inact ivated. We have thus far isolated Tc1 insertions in 16 different genes and obtained deletion derivatives of 6 of those.