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© Valérie Choumet
Mosquitoes were orally infected with the chikungunya virus. Midguts were dissected at day 5 post-infection, fixed and permeabilised. Virus is shown in red (anti-E2 protein, cyanine 3), the actin network in green (phalloidin 548) and nuclei in blue (DAPI).
Publication : Nature protocols

Efficient ΦC31 integrase-mediated site-specific germline transformation of Anopheles gambiae

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in Nature protocols - 19 Jun 2014

Pondeville E, Puchot N, Meredith JM, Lynd A, Vernick KD, Lycett GJ, Eggleston P, Bourgouin C

Link to Pubmed [PMID] – 24945385

Nat Protoc 2014 Jul;9(7):1698-712

Current transgenic methodology developed for mosquitoes has not been applied widely to the major malaria vector Anopheles gambiae, which has proved more difficult to genetically manipulate than other mosquito species and dipteran insects. In this protocol, we describe ΦC31-mediated site-specific integration of transgenes into the genome of A. gambiae. The ΦC31 system has many advantages over ‘classical’ transposon-mediated germline transformation systems, because it allows integration of large transgenes at specific, characterized genomic locations. Starting from a general protocol, we have optimized steps from embryo collection to co-injection of transgene-containing plasmid and in vitro-produced ΦC31 integrase mRNA. We also provide tips for screening transgenic larvae. The outlined procedure provides robust transformation in A. gambiae, resulting in homozygous transgenic lines in ∼2-3 months.

http://www.ncbi.nlm.nih.gov/pubmed/24945385