Drawing a high-resolution functional map of adeno-associated virus capsid by massively parallel sequencing

Kei Adachi, Tatsuji Enoki, Yasuhiro Kawano, Michael Veraz and Hiroyuki Nakai ()
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Kei Adachi: Oregon Health and Science University School of Medicine
Tatsuji Enoki: Takara Bio Inc.
Yasuhiro Kawano: Oregon Health and Science University School of Medicine
Michael Veraz: Oregon Health and Science University School of Medicine
Hiroyuki Nakai: Oregon Health and Science University School of Medicine

Nature Communications, 2014, vol. 5, issue 1, 1-14

Abstract: Abstract Adeno-associated virus (AAV) capsid engineering is an emerging approach to advance gene therapy. However, a systematic analysis on how each capsid amino acid contributes to multiple functions remains challenging. Here we show proof-of-principle and successful application of a novel approach, termed AAV Barcode-Seq, that allows us to characterize phenotypes of hundreds of different AAV strains in a high-throughput manner and therefore overcomes technical difficulties in the systematic analysis. In this approach, we generate DNA barcode-tagged AAV libraries and determine a spectrum of phenotypes of each AAV strain by Illumina barcode sequencing. By applying this method to AAV capsid mutant libraries tagged with DNA barcodes, we can draw a high-resolution map of AAV capsid amino acids important for the structural integrity and functions including receptor binding, tropism, neutralization and blood clearance. Thus, Barcode-Seq provides a new tool to generate a valuable resource for virus and gene therapy research.

Date: 2014
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DOI: 10.1038/ncomms4075

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