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Originally published as Genetics Published Articles Ahead of Print on March 23, 2009.
Genetics, Vol. 182, 25-32, May 2009, Copyright © 2009
doi:10.1534/genetics.109.101998
Identification of EMS-Induced Mutations in Drosophila melanogaster by Whole-Genome Sequencing
Justin P. Blumenstiel*,
,
Aaron C. Noll,
Jennifer A. Griffiths,
Anoja G. Perera,
Kendra N. Walton,
William D. Gilliland,
R. Scott Hawley,
and
Karen Staehling-Hampton,1
* Department of Ecology and Evolutionary Biology, University of Kansas, Lawrence, Kansas 66045, Stowers Institute for Medical Research, Kansas City, Missouri 64110 and Department of Physiology, Kansas University Medical Center, Kansas City, Kansas 66160
1 Corresponding Author: Stowers Institute for Medical Research, 1000 E. 50th St., Kansas City, MO 64110.
E-mail: ksh{at}stowers.org
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 
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.
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Genetics 2009 182: NP.
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