摘要
目的构建人肿瘤转移抑制基因ＮＭ２３－Ｈ１与真核表达载体ＰｃＤＮＡ３．１／Ｚｅｒｏ的重组体。方法利用分子生物学克隆技术从质粒ｐｕｃ１８中酶切回收９００ｂｐ的人ＮＭ２３－Ｈ１基因片段，与ＰｃＤＮＡ３．１／Ｚｅｒｏ构成重组体，并根据表达载体与克隆基因核苷酸序列中酶切位点分析、鉴定插入片段的正反连接。结果酶切得到９００ｂｐ的ＮＭ２３－Ｈ１基因片段，５．０ｋｂ质粒载体，两者相连构建成重组体ＰｃＤＮＡ．ＮＭ２３，利用ＰｓｔⅠ酶切位点确认ＰｓｔＩ酶切后得到４条带的重组体为正向连接。结论人真核细胞表达质粒ＰｃＤＮＡ．ＮＭ２３成功构建，为卵巢癌基因治疗提供了部分实验基础。
Objective\ To develop a somatic gene therapy system for ovarian carcinoma treatment, we constructed a human non metastatic gene(NM23 H1) expressing vector in ovarian carcinoma cells. Methods\ PUC18 NM23 H1 was treated with BamH1 to release the 900bp fragment. PcDNA3 1/Zero was digested with BamH1, releasing a 5 0kb fragment, NM23 H1(0 9kb) fragment was ligated to the vector PcDNA3 1/Zero(5 0kb). Results\ 900bp fragment and 5 0kb vector obtained by BamH1 digesting were connected with T4 DNA ligase to produce recombinant PcDNA·NM23. According to the enzymatic site PstⅠ, the positive oriention of the recombinant could be digested into four different fragments by PstⅠ. Conclusion\ The successful construction of PcDNA·NM could provide a rational basis for the gene therapy of ovarian carcinoma.
出处
《解剖学报》
CAS
CSCD
北大核心
1999年第2期171-173,共3页
Acta Anatomica Sinica
