Institut for Biomedicin

Lars Bolund

Efficient construction of rAAV-based gene targeting vectors by Golden Gate cloning

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The recombinant adeno-associated virus (rAAV) has proven to be an efficient and attractive tool for targeted genome engineering. Here we present a novel method employing the Golden Gate cloning strategy for fast and efficient construction of rAAV-based gene knockout or single-nucleotide knockin vectors. Two vectors, pGolden-Neo and pGolden-Hyg, were generated as common assembling modules to confer antibiotic resistance to the targeting vector. To validate the method, we then generated two rAAV-based targeting vectors: pAAV-pTP53-KO and pAAV-hTauP301L-KI. Furthermore, we generated a pGolden-AAV plasmid that allows one-step generation of an rAAV-based targeting vector. Our new methodology for rAAV targeting vector assembly is efficient, accurate, time-saving, and cost-effective.

OriginalsprogEngelsk
TidsskriftBioTechniques
Vol/bind56
Nummer5
Sider (fra-til)263-8
Antal sider6
ISSN0736-6205
DOI
StatusUdgivet - maj 2014

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