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CRISPR-Cas9-Guided Genome Engineering in Caenorhabditis elegans.

Publication ,  Journal Article
Kim, H-M; Colaiácovo, MP
Published in: Current protocols in molecular biology
December 2019

The CRISPR-Cas (clustered regularly interspaced short palindromic repeats-CRISPR-associated protein) system is being used successfully for efficient and targeted genome editing in various organisms, including the nematode Caenorhabditis elegans. Recent studies have developed a variety of CRISPR-Cas9 approaches to enhance genome engineering via two major DNA double-strand break repair pathways: nonhomologous end joining and homologous recombination. Here, we describe a protocol for Cas9-mediated C. elegans genome editing together with single guide RNA (sgRNA) and repair template cloning (canonical marker-free and cassette selection methods), as well as injection methods required for delivering Cas9, sgRNAs, and repair template DNA into the germline. © 2019 by John Wiley & Sons, Inc. Basic Protocol 1: Guide RNA preparation Alternate Protocol 1: sgRNA cloning using fusion PCR Basic Protocol 2: Preparation of a repair template for homologous recombination Alternate Protocol 2: Preparation of repair template donors for the cassette selection method Basic Protocol 3: Injecting animals Basic Protocol 4: Screening transgenic worms with marker-free method Alternate Protocol 3: Screening transgenic worms with cassette selection method.

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Published In

Current protocols in molecular biology

DOI

EISSN

1934-3647

ISSN

1934-3639

Publication Date

December 2019

Volume

129

Issue

1

Start / End Page

e106

Related Subject Headings

  • RNA, Guide, CRISPR-Cas Systems
  • Genetic Engineering
  • Gene Targeting
  • Gene Editing
  • Caenorhabditis elegans
  • CRISPR-Cas Systems
  • Biochemistry & Molecular Biology
  • Animals, Genetically Modified
  • Animals
 

Citation

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Chicago
ICMJE
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Kim, H.-M., & Colaiácovo, M. P. (2019). CRISPR-Cas9-Guided Genome Engineering in Caenorhabditis elegans. Current Protocols in Molecular Biology, 129(1), e106. https://doi.org/10.1002/cpmb.106
Kim, Hyun-Min, and Monica P. Colaiácovo. “CRISPR-Cas9-Guided Genome Engineering in Caenorhabditis elegans.Current Protocols in Molecular Biology 129, no. 1 (December 2019): e106. https://doi.org/10.1002/cpmb.106.
Kim H-M, Colaiácovo MP. CRISPR-Cas9-Guided Genome Engineering in Caenorhabditis elegans. Current protocols in molecular biology. 2019 Dec;129(1):e106.
Kim, Hyun-Min, and Monica P. Colaiácovo. “CRISPR-Cas9-Guided Genome Engineering in Caenorhabditis elegans.Current Protocols in Molecular Biology, vol. 129, no. 1, Dec. 2019, p. e106. Epmc, doi:10.1002/cpmb.106.
Kim H-M, Colaiácovo MP. CRISPR-Cas9-Guided Genome Engineering in Caenorhabditis elegans. Current protocols in molecular biology. 2019 Dec;129(1):e106.

Published In

Current protocols in molecular biology

DOI

EISSN

1934-3647

ISSN

1934-3639

Publication Date

December 2019

Volume

129

Issue

1

Start / End Page

e106

Related Subject Headings

  • RNA, Guide, CRISPR-Cas Systems
  • Genetic Engineering
  • Gene Targeting
  • Gene Editing
  • Caenorhabditis elegans
  • CRISPR-Cas Systems
  • Biochemistry & Molecular Biology
  • Animals, Genetically Modified
  • Animals