Quantitative comparison of expression with adeno-associated virus (AAV-2) brain-specific gene cassettes

R. Xu, C. G. Janson, M. Mastakov, P. Lawlor, D. Young, A. Mouravlev, H. Fitzsimons, K. L. Choi, H. Ma, M. Dragunow, P. Leone, Q. Chen, B. Dicker, M. J. During

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144 Scopus citations


This study compared a range of mammalian CNS expression cassettes in recombinant adeno-associated virus (AAV-2) vectors using strong endogenous promoter sequences, with or without a strong post-regulatory element and polyadenylation signal. Changes in these elements led to transgene expression varying by over three orders of magnitude. In experiments conducted in primary cell culture and in > 100 stereotactically injected rats, we observed highly efficient and stable (> 15 months) gene expression in neurons and limited expression in glia; the highest expression occurred with endogenous, nonviral promoters such as neuron-specific enolase and β-actin. The packaging size of AAV-2 was maximized at 5.7 kb without impairing gene expression, as judged by direct comparison with a number of smaller AAV-2 constructs. The genomic insert size and titer were confirmed by Southern blot and quantitative PCR, and infectivity was tested by particle titer using ELISA with a conformation-dependent epitope that requires the full intact capsid. A packaging and purification protocol we describe allows for high-titer, high-capacity AAV-2 vectors that can transduce over 2 × 105 neurons in vivo per microliter of vector, using the strongest expression cassette.

Original languageEnglish (US)
Pages (from-to)1323-1332
Number of pages10
JournalGene therapy
Issue number17
StatePublished - 2001
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Genetics
  • Molecular Medicine
  • Molecular Biology


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