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C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy

Whole-genome sequencing (WGS) is becoming a fast and cost-effective method to pinpoint molecular lesions in mutagenized genetic model systems, such as Caenorhabditis elegans. As mutagenized strains contain a significant mutational load, it is often still necessary to map mutations to a chromosomal i...

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Autores principales: Doitsidou, Maria, Poole, Richard J., Sarin, Sumeet, Bigelow, Henry, Hobert, Oliver
Formato: Texto
Lenguaje:English
Publicado: Public Library of Science 2010
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2975709/
https://www.ncbi.nlm.nih.gov/pubmed/21079745
http://dx.doi.org/10.1371/journal.pone.0015435
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author Doitsidou, Maria
Poole, Richard J.
Sarin, Sumeet
Bigelow, Henry
Hobert, Oliver
author_facet Doitsidou, Maria
Poole, Richard J.
Sarin, Sumeet
Bigelow, Henry
Hobert, Oliver
author_sort Doitsidou, Maria
collection PubMed
description Whole-genome sequencing (WGS) is becoming a fast and cost-effective method to pinpoint molecular lesions in mutagenized genetic model systems, such as Caenorhabditis elegans. As mutagenized strains contain a significant mutational load, it is often still necessary to map mutations to a chromosomal interval to elucidate which of the WGS-identified sequence variants is the phenotype-causing one. We describe here our experience in setting up and testing a simple strategy that incorporates a rapid SNP-based mapping step into the WGS procedure. In this strategy, a mutant retrieved from a genetic screen is crossed with a polymorphic C. elegans strain, individual F2 progeny from this cross is selected for the mutant phenotype, the progeny of these F2 animals are pooled and then whole-genome-sequenced. The density of polymorphic SNP markers is decreased in the region of the phenotype-causing sequence variant and therefore enables its identification in the WGS data. As a proof of principle, we use this strategy to identify the molecular lesion in a mutant strain that produces an excess of dopaminergic neurons. We find that the molecular lesion resides in the Pax-6/Eyeless ortholog vab-3. The strategy described here will further reduce the time between mutant isolation and identification of the molecular lesion.
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spelling pubmed-29757092010-11-15 C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy Doitsidou, Maria Poole, Richard J. Sarin, Sumeet Bigelow, Henry Hobert, Oliver PLoS One Research Article Whole-genome sequencing (WGS) is becoming a fast and cost-effective method to pinpoint molecular lesions in mutagenized genetic model systems, such as Caenorhabditis elegans. As mutagenized strains contain a significant mutational load, it is often still necessary to map mutations to a chromosomal interval to elucidate which of the WGS-identified sequence variants is the phenotype-causing one. We describe here our experience in setting up and testing a simple strategy that incorporates a rapid SNP-based mapping step into the WGS procedure. In this strategy, a mutant retrieved from a genetic screen is crossed with a polymorphic C. elegans strain, individual F2 progeny from this cross is selected for the mutant phenotype, the progeny of these F2 animals are pooled and then whole-genome-sequenced. The density of polymorphic SNP markers is decreased in the region of the phenotype-causing sequence variant and therefore enables its identification in the WGS data. As a proof of principle, we use this strategy to identify the molecular lesion in a mutant strain that produces an excess of dopaminergic neurons. We find that the molecular lesion resides in the Pax-6/Eyeless ortholog vab-3. The strategy described here will further reduce the time between mutant isolation and identification of the molecular lesion. Public Library of Science 2010-11-08 /pmc/articles/PMC2975709/ /pubmed/21079745 http://dx.doi.org/10.1371/journal.pone.0015435 Text en Doitsidou et al. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
spellingShingle Research Article
Doitsidou, Maria
Poole, Richard J.
Sarin, Sumeet
Bigelow, Henry
Hobert, Oliver
C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy
title C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy
title_full C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy
title_fullStr C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy
title_full_unstemmed C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy
title_short C. elegans Mutant Identification with a One-Step Whole-Genome-Sequencing and SNP Mapping Strategy
title_sort c. elegans mutant identification with a one-step whole-genome-sequencing and snp mapping strategy
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2975709/
https://www.ncbi.nlm.nih.gov/pubmed/21079745
http://dx.doi.org/10.1371/journal.pone.0015435
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