血管形成抑制剂Tumstatin在鼻咽癌中表达鉴定及基因克隆

Identification of the Expression of Anti-angiogenic Factor:Tumstatin in NPC and Gene Clone

目的:检测Tumstatin基因在鼻咽癌发病过程中的表达变化及克隆其编码序列。方法:利用半定量RT-PCR方法检测Tumstatin在鼻咽癌组织、鼻咽癌细胞和正常鼻咽组织中的表达,比较它们之间的表达差异。设计含酶切位点的PCR引物,利用RT-PCR方法从相对正常鼻咽组织中获取Tumstatin蛋白编码序列,T/A克隆入pMD18载体中。利用PCR和酶切鉴定获得阳性重组子。重组子最后经测序证实。结果:与相对正常鼻咽组织相比,Tumstatin在鼻咽癌组织和细胞中表达下调或缺失。RT-PCR法成功获得Tumstatin基因编码区全长cDNA序列。重组克隆质粒插入片段经DNA测序后与GenBank中Tumstatin基因相应序列比较,100%同源。结论:Tumstatin在鼻咽癌组织和细胞中表达下调或缺失。采用T/A技术成功克隆鼻咽组织中Tumstatin基因,为下一步将Tumstatin亚克隆入真核表达载体pEGFP-N1,探索其在鼻咽癌生长和转移中的作用奠定了基础。

Objective To detect the expression alteration of tumstatin gene in NPC pathogenesis and clone the CDS sequence of tumstatin.Methods Semiquantitative RT-PCR was used to detect the expression differentiation of tumstatin among nasopharyngeal carcinoma(NPC) tissues,NPC cells and normal nasopharyngeal epithelium(NP).PCR primers with the design of containing restriction endonuclease sites were utilized to amplify CDS sequence of tumstatin from human normal NP.The PCR product was then inserted into pMD18 vector.T...