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Genetics. 2017 Oct;207(2):447-463. doi: 10.1534/genetics.117.300179. Epub 2017 Aug 21.

MIP-MAP: High-Throughput Mapping of Caenorhabditis elegans Temperature-Sensitive Mutants via Molecular Inversion Probes.

Author information

1
Department of Genome Sciences, University of Washington, Seattle, Washington 98195 mok.calvin@gmail.com.
2
Department of Zoology and Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada.
3
Department of Genome Sciences, University of Washington, Seattle, Washington 98195.
4
Institute of Molecular Biology, University of Oregon, Eugene, Oregon 97403.
5
Department Biology II, Biocenter, Ludwig-Maximilians-Universität München, 82152 Planneg-Martinsried, Germany.
6
Department of Molecular Biology and Genetics, Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
7
Horace Mann School, Bronx, New York 10471.
8
Department of Developmental Genetics, Institute of Genetics, Technische Universität Braunschweig, 38106, Germany.

Abstract

Mutants remain a powerful means for dissecting gene function in model organisms such as Caenorhabditis elegans Massively parallel sequencing has simplified the detection of variants after mutagenesis but determining precisely which change is responsible for phenotypic perturbation remains a key step. Genetic mapping paradigms in C. elegans rely on bulk segregant populations produced by crosses with the problematic Hawaiian wild isolate and an excess of redundant information from whole-genome sequencing (WGS). To increase the repertoire of available mutants and to simplify identification of the causal change, we performed WGS on 173 temperature-sensitive (TS) lethal mutants and devised a novel mapping method. The mapping method uses molecular inversion probes (MIP-MAP) in a targeted sequencing approach to genetic mapping, and replaces the Hawaiian strain with a Million Mutation Project strain with high genomic and phenotypic similarity to the laboratory wild-type strain N2 We validated MIP-MAP on a subset of the TS mutants using a competitive selection approach to produce TS candidate mapping intervals with a mean size < 3 Mb. MIP-MAP successfully uses a non-Hawaiian mapping strain and multiplexed libraries are sequenced at a fraction of the cost of WGS mapping approaches. Our mapping results suggest that the collection of TS mutants contains a diverse library of TS alleles for genes essential to development and reproduction. MIP-MAP is a robust method to genetically map mutations in both viable and essential genes and should be adaptable to other organisms. It may also simplify tracking of individual genotypes within population mixtures.

KEYWORDS:

Caenorhabditis elegans; genetic mapping; massively multiplex sequencing; molecular inversion probes; temperature-sensitive mutations

PMID:
28827289
PMCID:
PMC5629315
DOI:
10.1534/genetics.117.300179
[Indexed for MEDLINE]
Free PMC Article

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