Nkx2.2的克隆及其在C6细胞中的表达  

Cloning and expression of Nkx2.2 in C6 cells

作　　者：王伟[1] 王涛[1] 徐清[1] 关路媛[1] 张斌[1] 

机构地区：[1]第四军医大学基础部生物化学与分子生物学教研室,陕西西安710033

出　　处：《第四军医大学学报》2009年第18期1651-1654,共4页Journal of the Fourth Military Medical University

基　　金：国家自然科学基金(30770420;30871239)

摘　　要：目的:克隆大鼠转录因子Nkx2.2,构建其真核表达载体,并观察其在大鼠C6胶质瘤细胞系中的表达.方法:利用RT-PCR从大鼠脑组织总RNA中扩增包含Nkx2.2编码区的cDNA片段,克隆连接至pMD20-T载体,再以pMD20-T-Nkx2.2为模板扩增出Nkx2.2编码区序列,将其克隆到真核表达载体pcDNA3.1-myc-his(-)中,测序验证后,转染C6胶质瘤细胞.用anti-His抗体和间接免疫荧光染色法检测Nkx2.2在C6细胞中的表达.结果:测序证实所克隆的Nkx2.2编码区正确地连接入pcDNA3.1-myc-his(-)中,免疫荧光检测证实其在C6细胞中得到了有效表达.结论:成功克隆了大鼠Nkx2.2编码区片段,构建了其真核表达载体,并在C6细胞中得到了有效表达,为进一步研究Nkx2.2基因的功能,尤其是探讨其在神经系统中的作用奠定了基础.AIM： To clone rat transcription factor Nkx2.2 and construct its eukaryotic expression vector, express it in C6 glioma cells. METHODS： RT-PCR was used to amplify fragment including Nkx2. 2 gene code region from rat brain RNA, the PCR product was inserted into pMD20-T vector, then amplified Nkx2, 2 coding region from pMD20-T-Nkx2. 2 and inserted it into eukaryotic expression vector pcDNA3.1-myc-his（ - ）. After being confirmed by sequencing, the pcDNA3.1-Nkx2.2 was transfected into C6 glioma ceils. The expression of Nkx2. 2 in C6 cells was detected by indirect immunofluorescent staining. RESULTS： Sequence analysis conformed that Nkx2. 2 was correctly inserted into pcDNA3. 1-myc-his （ - ） vector, C6 cells transfected with Nkx2. 2 could efficiently express Nkx2. 2 in cell nuclear. CONCLUSION： The cloning of rat Nkx2.2 and the construction of its eukaryotic expression vector were successful, Nkx2. 2 was efficiently expressed in C6 cell nuclear. This study would lay the foundation for further research on the function of Nkx2. 2 and its role in nervous system.

关 键 词：Nkx2.2 克隆 C6细胞 真核表达 

分 类 号：Q254[生物学—细胞生物学]