CORE--VECTOR FACILITY

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The Vector Core plays an integral and essential role in the production of research grade recombinant viral vectors for all of the investigators in the Program. It is critical that all investigators be provided with consistently high titer, pure rAAV preparations. It is particularly important that preps be free of replication-competent AAV (rcAAV) and have a particle to infectivity ratio of 200 or less. Therefore, the primary objective of the Vector Core is to make the highest quality vector possible for the investigators. Each virus preparation will be produced using a mini-Ad plasmid DNA system (to essentially eliminate wtAd contamination) and will be purified by a novel method utilizing iodixanol gradient centrifugation followed by heparin affinity chromatography. All virus stocks are subject to quality control assays to assess purity, particle titer, infectious titer, and potential contamination by rcAAV. The second objective of the Vector Core is to develop and implement improved methods in the process of scaling production and purification of rAAV. The Core is to develop and implement improved methods in the process of scaling production and purification of rAAV. The Vector Core is currently working on issues of scale-up production, including development of better helper systems. Additionally, we are testing new purification methods that include a variety of FPLC chromatography formats. Not only is the Vector Core able to make high quality rAAV virus preparations, but the Vector Core is also available to provide consulting on and/or actually perform vector construction and to aid in the development of biological assays is necessary for transgene screening. The Vector Core has already made available a variety of promoter-marker gene constructs for general screening.