Naoki Tamura, Kanzo Suzuki, Hirono Shiraki, Issei Waguri, Eri Segi-Nishida
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引用次数: 0
Abstract
Recombinant adeno-associated virus (AAV) vectors are widely used for manipulating gene expression. AAVrh.10 is a highly infectious AAV serotype for the central nervous system and various tissues. Owing to its potential use in research, we aimed to optimize the production strategy and develop a simple purification protocol for the AAVrh.10 vector. In this study, we explored a simple production and purification strategy for the AAVrh.10 vector via chloroform extraction and ultrafiltration. Initially, we evaluated the optimal conditions for AAVrh.10-CAG-GFP production using AAV-293 cells. AAVrh.10-CAG-GFP was successfully produced in a serum-free medium after plasmid transfection. Moreover, the culture medium contained a substantial amount of the virus. Therefore, both AAVrh.10-containing cell lysate and culture medium should be used to prepare the AAVrh.10 viral vector. To purify and concentrate AAVrh.10-CAG-GFP from the crude lysate and medium, we optimized the chloroform extraction and ultrafiltration strategies. Subsequently, purified AAVrh.10-CAG-GFP was used to infect HEK-293T cells. Overall, this study provides a simple and effective AAVrh.10 vector preparation strategy for basic and preclinical research.
期刊介绍:
Biological and Pharmaceutical Bulletin (Biol. Pharm. Bull.) began publication in 1978 as the Journal of Pharmacobio-Dynamics. It covers various biological topics in the pharmaceutical and health sciences. A fourth Society journal, the Journal of Health Science, was merged with Biol. Pharm. Bull. in 2012.
The main aim of the Society’s journals is to advance the pharmaceutical sciences with research reports, information exchange, and high-quality discussion. The average review time for articles submitted to the journals is around one month for first decision. The complete texts of all of the Society’s journals can be freely accessed through J-STAGE. The Society’s editorial committee hopes that the content of its journals will be useful to your research, and also invites you to submit your own work to the journals.