论文部分内容阅读
应用分子克隆技术构建了含８４３ｂｐ的全长人ＩＦＮ－γｃＤＮＡ的逆转录病毒表达载体Ｎ２／ＩＦＮ，采用磷酸钙共沉淀方法转染包装细胞ΨＣＲＥ和ΨＣＲＩＰ，并能得到较高滴度的含病毒颗粒上清。然后应用ΨＣＲＩＰ细胞培养上清的复制缺陷性病毒颗粒感染Ｌｅｗｉｓ肺癌细胞（ＬＬＣ），经过Ｇ４１８筛选克隆得到一株能稳定分泌ＩＦＮ（６４００Ｕ／Ｍ）的克隆，Ｓｏｕｔｈｅｒｎ印迹证实ＩＦＮ－γ基因已插入ＬＬＣ基因组。结果表明，该逆转录病毒载体系统能有效地介导基因转移，并能使目的基因在靶细胞稳定表达。结果为肿瘤基因治疗及制备新型肿瘤疫苗奠定了基础。
The retroviral expression vector N2 / IFN containing 843bp full-length human IFN-γcDNA was constructed by molecular cloning technique, and the packaging cells ΨCRE and ΨCRIP were transfected by the calcium phosphate coprecipitation method, and the higher titer of the virus-containing particles Supernatant. Then, Lewis lung cancer cells (LLC) were infected with replication-defective virus particles of ΨCRIP cell culture supernatant and cloned by G418 to obtain a clone capable of stably secreting IFN (6400U / M). The Southern blot confirmed that the IFN-γ gene was inserted into LLC Genome. The results showed that the retroviral vector system can effectively mediate gene transfer and make the target gene stably expressed in target cells. The results for the tumor gene therapy and preparation of new tumor vaccine laid the foundation.