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Genetics. 2009 May;182(1):25-32. doi: 10.1534/genetics.109.101998. Epub 2009 Mar 23.

Identification of EMS-induced mutations in Drosophila melanogaster by whole-genome sequencing.

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  • 1Department of Physiology, Kansas University Medical Center, Kansas City, Kansas 66160, USA.

Abstract

Next-generation methods for rapid whole-genome sequencing enable the identification of single-base-pair mutations in Drosophila by comparing a chromosome bearing a new mutation to the unmutagenized sequence. To validate this approach, we sought to identify the molecular lesion responsible for a recessive EMS-induced mutation affecting egg shell morphology by using Illumina next-generation sequencing. After obtaining sufficient sequence from larvae that were homozygous for either wild-type or mutant chromosomes, we obtained high-quality reads for base pairs composing approximately 70% of the third chromosome of both DNA samples. We verified 103 single-base-pair changes between the two chromosomes. Nine changes were nonsynonymous mutations and two were nonsense mutations. One nonsense mutation was in a gene, encore, whose mutations produce an egg shell phenotype also observed in progeny of homozygous mutant mothers. Complementation analysis revealed that the chromosome carried a new functional allele of encore, demonstrating that one round of next-generation sequencing can identify the causative lesion for a phenotype of interest. This new method of whole-genome sequencing represents great promise for mutant mapping in flies, potentially replacing conventional methods.

PMID:
19307605
[PubMed - indexed for MEDLINE]
PMCID:
PMC2674820
Free PMC Article

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