Format

Send to

Choose Destination
Nat Commun. 2014;5:3075. doi: 10.1038/ncomms4075.

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

Author information

1
1] Department of Molecular and Medical Genetics, Oregon Health and Science University School of Medicine, Portland, Oregon 97239, USA [2].
2
1] Takara Bio Inc., Otsu, Shiga 520-2134, Japan [2].
3
1] Department of Molecular and Medical Genetics, Oregon Health and Science University School of Medicine, Portland, Oregon 97239, USA [2] Takara Bio Inc., Otsu, Shiga 520-2134, Japan.
4
Department of Molecular and Medical Genetics, Oregon Health and Science University School of Medicine, Portland, Oregon 97239, USA.

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.

PMID:
24435020
PMCID:
PMC3941020
DOI:
10.1038/ncomms4075
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for Nature Publishing Group Icon for PubMed Central
Loading ...
Support Center