Site-selected mutagenesis of the Drosophila second chromosome via plasmid rescue of lethal P-element insertions

Yiquan Guo, Ann Gillan, Tibor Török, I. Kiss, Julian A T Dow, Kim Kaiser

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15 Citations (Scopus)

Abstract

This paper describes a fast and efficient approach to correlating cloned genes with mutant phenotypes in Drosophila. We make use of a large collection D. melanogaster lines with recessive lethal insertions of a P[lacW] transposon on their second chromosome. Within this collection there clearly must be many insertions corresponding to Drosophila genes that have been cloned and characterized, e.g., via homology with cloned mammalian genes, but for which mutant phenotypes have yet to be identified. We have made use of the fact that P[lacW] contains a plasmid replicon to establish a collection of rescued plasmids containing genomic DNA flanking the sites of transposon insertion. Plasmids representing a total of 1836 lines were independently rescued and pooled in batches of 10 and 100. Pools of 100 plasmids were screened by hybridization with cDNAs corresponding to cloned second chromosome loci. Hybridizing pools were then narrowed down to single plasmids by a process of subdivision and rehybridization, and corresponding mutant lines were obtained. The success rate was better than one in four. This rate would undoubtedly be improved by the use of genomic DNA probes.

Original languageEnglish
Pages (from-to)972-979
Number of pages8
JournalGenome Research
Volume6
Issue number10
Publication statusPublished - 1996

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Mutagenesis
Drosophila
Plasmids
Chromosomes
Genes
Phenotype
Replicon
DNA Probes
Complementary DNA
DNA

ASJC Scopus subject areas

  • Genetics

Cite this

Site-selected mutagenesis of the Drosophila second chromosome via plasmid rescue of lethal P-element insertions. / Guo, Yiquan; Gillan, Ann; Török, Tibor; Kiss, I.; Dow, Julian A T; Kaiser, Kim.

In: Genome Research, Vol. 6, No. 10, 1996, p. 972-979.

Research output: Contribution to journalArticle

Guo, Y, Gillan, A, Török, T, Kiss, I, Dow, JAT & Kaiser, K 1996, 'Site-selected mutagenesis of the Drosophila second chromosome via plasmid rescue of lethal P-element insertions', Genome Research, vol. 6, no. 10, pp. 972-979.
Guo, Yiquan ; Gillan, Ann ; Török, Tibor ; Kiss, I. ; Dow, Julian A T ; Kaiser, Kim. / Site-selected mutagenesis of the Drosophila second chromosome via plasmid rescue of lethal P-element insertions. In: Genome Research. 1996 ; Vol. 6, No. 10. pp. 972-979.
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