Design and construction of targeted AAVP vectors for mammalian cell transduction

Amin Hajitou, Roberto Rangel, Martin Trepel, Suren Soghomonyan, Juri G. Gelovani, Mian M. Alauddin, Renata Pasqualini, Wadih Arap

Research output: Contribution to journalArticlepeer-review

81 Scopus citations

Abstract

Bacteriophage (phage) evolved as bacterial viruses, but can be adapted to transduce mammalian cells through ligand-directed targeting to a specific receptor. We have recently reported a new generation of hybrid prokaryotic-eukaryotic vectors, which are chimeras of genetic cis-elements of recombinant adeno-associated virus and phage (termed AAVP). This protocol describes the design and construction of ligand-directed AAVP vectors, production of AAVP particles and the methodology to transduce mammalian cells in vitro and to target tissues in vivo after systemic administration. Targeted AAVP particles are made in a two-step process. First, a ligand peptide of choice is displayed on the coat protein to generate a targeted backbone phage vector. Then, a recombinant AAV carrying a mammalian transgene cassette is inserted into an intergenomic region. High-titer suspensions (∼1010 -1011 transducing units per μl) can be produced within 3 days after vector construction. Transgene expression by targeted AAVP usually reaches maximum levels within 1 week.

Original languageEnglish (US)
Pages (from-to)523-531
Number of pages9
JournalNature Protocols
Volume2
Issue number3
DOIs
StatePublished - Mar 2007
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry, Genetics and Molecular Biology(all)

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