论文部分内容阅读
目的:构建针对人鼻咽癌CNE-2Z细胞Bcl-2基因pENTR-CMV-EGFP-hsa-mir-16-1/15a真核表达质粒,转染至CNE-2Z细胞并检测其表达。方法:采用PCR法从重组质粒PGH-16-1/15a中获得16-1/15a-X2370G全长序列,在T4DNALigase连接酶作用下连接入重组载体pENTR-CMV-EGFP。重组质粒经酶切及测序鉴定。将构建成功的重组质粒转染入人鼻咽癌细胞株CNE-2Z,用荧光显微镜观察转染结果。结果:重组质粒pENTR-CMV-EGFP-hsa-mir-16-1/15a经酶切与测序证实构建成功,转染至鼻咽癌CNE-2Z细胞后,荧光显微镜观察证实该重组质粒能在CNE-2Z中表达。结论:成功构建真核表达质粒pENTR-CMV-EGFP-hsa-mir-16-1/15a,并在鼻咽癌CNE-2Z细胞中得到表达,可用于进一步检测其抗肿瘤机制。
OBJECTIVE: To construct the eukaryotic expression vector pENTR-CMV-EGFP-hsa-mir-16-1 / 15a targeting human Bcl-2 gene of human nasopharyngeal carcinoma CNE-2Z cells and transfect it into CNE-2Z cells for detection of their expression. METHODS: The full-length sequence of 16-1 / 15a-X2370G was obtained from the recombinant plasmid PGH-16-1 / 15a by PCR and ligated into the recombinant vector pENTR-CMV-EGFP under the action of T4 DNase ligase. Recombinant plasmids were identified by restriction enzyme digestion and sequencing. The constructed recombinant plasmids were transfected into human nasopharyngeal carcinoma cell line CNE-2Z, and the transfection results were observed under a fluorescence microscope. Results: The recombinant plasmid pENTR-CMV-EGFP-hsa-mir-16-1 / 15a was confirmed by restriction enzyme digestion and sequencing. After transfection into CNE-2Z cells, the recombinant plasmid pEGFP- -2Z expression. CONCLUSION: The eukaryotic expression plasmid pENTR-CMV-EGFP-hsa-mir-16-1 / 15a has been successfully constructed and expressed in CNE-2Z cells, which can be used to further detect its anti-tumor mechanism.