Advanced

AAV Production Everywhere : A Simple, Fast, and Reliable Protocol for In-house AAV Vector Production Based on Chloroform Extraction

Negrini, Matilde LU ; Wang, Gang LU ; Heuer, Andreas LU ; Björklund, Tomas LU and Davidsson, Marcus LU (2020) In Current Protocols in Neuroscience 93(1). p.103-103
Abstract

Recombinant adeno-associated virus (rAAV) is a mammalian virus that has been altered to be used as a gene delivery vehicle. Several changes to the viral genome have made them replication deficient so that this aspect of the viral infection cycle is under full control of the experimenter, while maintaining gene expression machinery. Over the last decades, rAAVs have become the gold standard for studying in vivo gene function and are especially favorable for gene transfer in the central nervous system. AAVs have been proven safe and provide stable gene expression over a long period of time. They are extensively used in preclinical experiments and show great potential for clinical applications. However, the use of AAVs in preclinical... (More)

Recombinant adeno-associated virus (rAAV) is a mammalian virus that has been altered to be used as a gene delivery vehicle. Several changes to the viral genome have made them replication deficient so that this aspect of the viral infection cycle is under full control of the experimenter, while maintaining gene expression machinery. Over the last decades, rAAVs have become the gold standard for studying in vivo gene function and are especially favorable for gene transfer in the central nervous system. AAVs have been proven safe and provide stable gene expression over a long period of time. They are extensively used in preclinical experiments and show great potential for clinical applications. However, the use of AAVs in preclinical settings are often held back due to availability. Waiting lines are long at commercial production facilities, and in-lab production is hindered due to lack of specific laboratory equipment needed. Here we present a novel production method that can be carried out in any molecular biology laboratory using standard laboratory equipment. We provide a simple, fast, and streamlined protocol for production that can result in titers comparable with the more time-consuming iodixanol gradient ultracentrifugation method. The yield using this protocol is high enough for any type of study where AAV is the vector of choice.

(Less)
Please use this url to cite or link to this publication:
author
; ; ; and
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
AAV vector, adeno associated virus, chloroform extraction, gene expression, gene therapy, PEI transfection, vector production
in
Current Protocols in Neuroscience
volume
93
issue
1
pages
103 - 103
publisher
John Wiley and Sons Inc.
external identifiers
  • pmid:32865885
  • scopus:85090106089
ISSN
1934-8584
DOI
10.1002/cpns.103
language
English
LU publication?
yes
id
4c9fad9f-db7c-4439-ac1a-13cc5d29928d
date added to LUP
2020-09-25 10:02:52
date last changed
2020-09-27 01:59:51
@article{4c9fad9f-db7c-4439-ac1a-13cc5d29928d,
  abstract     = {<p>Recombinant adeno-associated virus (rAAV) is a mammalian virus that has been altered to be used as a gene delivery vehicle. Several changes to the viral genome have made them replication deficient so that this aspect of the viral infection cycle is under full control of the experimenter, while maintaining gene expression machinery. Over the last decades, rAAVs have become the gold standard for studying in vivo gene function and are especially favorable for gene transfer in the central nervous system. AAVs have been proven safe and provide stable gene expression over a long period of time. They are extensively used in preclinical experiments and show great potential for clinical applications. However, the use of AAVs in preclinical settings are often held back due to availability. Waiting lines are long at commercial production facilities, and in-lab production is hindered due to lack of specific laboratory equipment needed. Here we present a novel production method that can be carried out in any molecular biology laboratory using standard laboratory equipment. We provide a simple, fast, and streamlined protocol for production that can result in titers comparable with the more time-consuming iodixanol gradient ultracentrifugation method. The yield using this protocol is high enough for any type of study where AAV is the vector of choice.</p>},
  author       = {Negrini, Matilde and Wang, Gang and Heuer, Andreas and Björklund, Tomas and Davidsson, Marcus},
  issn         = {1934-8584},
  language     = {eng},
  number       = {1},
  pages        = {103--103},
  publisher    = {John Wiley and Sons Inc.},
  series       = {Current Protocols in Neuroscience},
  title        = {AAV Production Everywhere : A Simple, Fast, and Reliable Protocol for In-house AAV Vector Production Based on Chloroform Extraction},
  url          = {http://dx.doi.org/10.1002/cpns.103},
  doi          = {10.1002/cpns.103},
  volume       = {93},
  year         = {2020},
}