Journal
G3-GENES GENOMES GENETICS
Volume 8, Issue 2, Pages 653-658Publisher
GENETICS SOCIETY AMERICA
DOI: 10.1534/g3.117.1134
Keywords
Anopheles; gene drive; reverse genetics; transgenics; CRISPR; Cas9
Categories
Funding
- National Institutes of Health (NIH) [1R01 AI-113248, 1R21 AI-115271]
- NIH [5K22 AI-113060, 1R21 AI-123937]
- Defense Advanced Research Project Agency Safe Genes Program [HR0011-17-2-0047]
- NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES [K22AI113060, R21AI123937, R01AI113248, R21AI115271] Funding Source: NIH RePORTER
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Anopheles mosquitoes transmit at least 200 million annual malaria infections worldwide. Despite considerable genomic resources, mechanistic understanding of biological processes in Anopheles has been hampered by a lack of tools for reverse genetics. Here, we report successful application of the CRISPR/Cas9 system for highly efficient, site-specific mutagenesis in the diverse malaria vectors Anopheles albimanus, A. coluzzii, and A. funestus. When guide RNAs (gRNAs) and Cas9 protein are injected at high concentration, germline mutations are common and usually biallelic, allowing for the rapid creation of stable mutant lines for reverse genetic analysis. Our protocol should enable researchers to dissect the molecular and cellular basis of anopheline traits critical to successful disease transmission, potentially exposing new targets for malaria control.
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