CORE--VECTOR FACILITY

载体核心在生产研究级重组病毒载体中发挥着不可或缺的重要作用。为所有研究人员提供一贯高滴度、纯净的rAAV制剂是至关重要的。尤其重要的是，制备物不含具有复制能力的AAV (rcAAV)，并且颗粒感染性比小于等于200。因此，Vector Core的主要目标是为研究者提供最高质量的Vector。每种病毒制剂将使用mini-Ad质粒DNA系统（基本上消除wtAd污染）生产，并将通过利用碘二醇梯度离心和肝素亲和层析的新方法纯化。所有病毒储存库都要进行质量控制分析，以评估纯度、颗粒滴度、感染滴度和rcAAV的潜在污染。Vector Core的第二个目标是开发和实施rAAV规模化生产和纯化过程中的改进方法。核心是开发和实施rAAV规模化生产和纯化过程中的改进方法。Vector Core目前正在研究规模化生产的问题，包括开发更好的辅助系统。此外，我们正在测试新的净化方法，包括各种FPLC色谱格式。Vector Core不仅能够制作高质量的rAAV病毒制剂，而且还可以提供咨询和/或实际进行载体构建，并帮助开发转基因筛选所需的生物检测方法。Vector Core已经为一般筛选提供了多种启动子标记基因结构。

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.