A Robust and All-Inclusive Pipeline for Shuffling of Adeno-Associated Viruses.
ACS Synth Biol
; 8(1): 194-206, 2019 01 18.
Article
em En
| MEDLINE
| ID: mdl-30513195
ABSTRACT
Adeno-associated viruses (AAV) are attractive templates for engineering of synthetic gene delivery vectors. A particularly powerful technology for breeding of novel vectors with improved properties is DNA family shuffling, i.e., generation of chimeric capsids by homology-driven DNA recombination. Here, to make AAV DNA shuffling available to a wider community, we present a robust experimental and bioinformatical pipeline comprising (i) standardized and partially codon-optimized plasmids carrying 12 different AAV capsid genes; (ii) a scalable protocol including troubleshooting guide for viral library production; and (iii) the freely available software SALANTO for comprehensive analysis of chimeric AAV DNA and protein sequences. Moreover, we describe a set of 12 premade and ready-to-use AAV libraries. Finally, we demonstrate the usefulness of DNA barcoding technology to trace AAV capsid libraries within a complex mixture. Our protocols and resources facilitate the implementation and tailoring of AAV evolution technology in any laboratory interested in customized viral gene transfer.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Dependovirus
/
Vetores Genéticos
Tipo de estudo:
Risk_factors_studies
Idioma:
En
Ano de publicação:
2019
Tipo de documento:
Article