CONSTRUCTION OF ADENOVIRUS VECTORS

腺病毒载体介导的基因转移尤其引人注目 对于标准转染方法难治的细胞， 例如磷酸钙沉淀、脂质体或 电穿孔 这包括原代细胞培养和其他 高度分化的细胞，包括非分裂细胞。 为 出于同样的原因，腺病毒载体是选择的方法， 原位基因转移 此外，腺病毒载体可用于 在细胞中同时或同时表达几种外源基因， 以规定的时间间隔。 基因表达的水平是可以控制的 使用的病毒剂量。 该项目的目的是 构建用于体外使用的复制缺陷型载体。 这些 载体将指导多种蛋白质的表达， 是感兴趣的研究人员在NIDR部门的校内 Research. 这些载体将用于研究信号转导， 细胞外基质和唾液腺生理学。 具体而言是 合同的第一个目标是培养和纯化质粒 含有腺病毒序列。 第二个将是 重组质粒以产生腺病毒载体。 第三个目标 该项目的主要目的是扩增分离的腺病毒噬斑， 将其纯化成病毒原液。 这些试剂将是独特的， 无法从商业来源获得。

Adenoviral vector-mediated gene transfer is particularly attractive for cells that are refractory to standard methods of transfection, such as calcium phosphate precipitation, liposomes or electroporation. This includes primary cell cultures and other highly differentiated cells, including non-dividing cells. For the same reasons, adenoviral vectors are the method of choice for in situ gene transfer. In addition, adenoviral vectors may be used to express several exogenous genes in a cell, either simultaneously or at defined intervals. Levels of gene expression can be controlled by the dose of virus used. The purpose of this project is to construct replication-deficient vectors for in vitro use. These vectors will direct the expression of a wide range of proteins that are of interest to investigators in the NIDR Division of Intramural Research. The vectors will be used to study signal transduction, extracellular matrix and salivary gland physiology. Specifically, the first goal of the contract will be to grow and purify plasmids containing adenovirus sequences. The second will be to recombine plasmids to generate adenovirus vectors. The third goal of the project will be to amplify isolated adenovirus plaques and to purify them into virus stocks. These reagents will be unique and not available from commercial sources.