Construction and characterization of deletions with defined end points in Drosophila using P elements in trans.
We used P-element transposase-mediated "male recombination" between two P elements in trans to create genetic deletions that removed a number of loci, including the gene encoding the neuropeptide crustacean cardioactive peptide (CCAP). Two classes of recombinant chromosomes were produced. Approximately one-quarter were viable when homozygous or hemizygous, whereas the remaining lines caused homozygous and hemizygous lethality. Preliminary analyses using PCR and CCAP immunohistochemistry suggested that, whereas the DNA of the viable lines was largely intact, most lethal lines contained chromosomal deletions that were roughly bounded by the insertion sites of the two P elements used. Southern blot analyses of select lethal lines showed that the DNA flanking the deletion was indeed grossly intact whereas the intervening DNA could not be detected. Sequencing across the deletion in three of these lethal lines identified a single line bearing intact genomic DNA on either side of the deletion separated by 30 bp of P-element DNA. The method described here suggests a simple procedure for creating deletions with defined end points. Importantly, it can use preexisting P-element insertion strains and does not rely on the use of transposable elements that are engineered to cause specific DNA rearrangements.
Paré
Adam C
AC
Entomology Department, Cornell University, Ithaca, New York 14853, USA.
Dean
Derek M
DM
Ewer
John
J
D000818:Animals; D015139:Blotting, Southern; D002875:Chromosomes / Q000235:genetics; D004251:DNA Transposable Elements / Q000235:genetics*; D004331:Drosophila melanogaster / Q000235:genetics*; D017353:Gene Deletion; D005818:Genetic Engineering / Q000379:methods*; D006720:Homozygote; D007814:Larva / Q000166:cytology; D008297:Male; D016296:Mutagenesis; D009479:Neuropeptides / Q000235:genetics; D011995:Recombination, Genetic / Q000235:genetics; D017422:Sequence Analysis, DNA
D004251:DNA Transposable Elements; D009479:Neuropeptides; C058620:crustacean cardioactive peptide