摘要
目的构建pc DNATM6.2-GW-miR-22表达载体,并检测其对结直肠癌HCT116细胞增殖的影响。方法人工合成pre-hsa-miR-22基因片段,应用分子克隆技术构建pc DNATM6.2-GW-miR-22表达载体,酶切、DNA测序及BLAST验证;瞬时转染HCT116细胞,应用RT-PCR检测miR-22表达水平,MTT实验检测miR-22对HCT116细胞增殖的影响。结果酶切及测序结果验证miR-22表达载体构建成功;转染HCT116细胞后miR-22表达水平显著增高;MTT结果表明miR-22对细胞增殖具有抑制作用。结论本研究成功构建miR-22真核表达载体,证明miR-22高表达可以抑制结直肠癌细胞HCT116的增殖。
Objective To construct pcDNATM6.2-GW-miR-22 expression vector, and to explore its effect on the proliferation of color- ectal cancer cell line HCT116. Methods The pre-hsa-miR-22 gene fragments were artificially synthesized. The expression vector pcDNATM6.2-GW-miR-22 was constructed by molecular cloning technique and then verified by restriction enzyme digestion, sequence alignment and BLAST. The vector was transfected into HCT116 cells, and the expression level was determined by RT-PCR. The prolif- eration of HCT116 ceils was examined by M3~F assay. Results The pcDNATM6.2-GW-miR-22 expression vector was successfully constructed and confirmed by the results of restriction enzyme digestion and sequencing. The expression level of miR-22 in HCT-116 cells transfected with expression vectors was increased significantly（ P 〈0.05 ）. MTT assay showed that miR-22 inhibited the prolifera- tion of HCT116 cells. Conclusion The pcDNATM6.2-GW-miR-22 expression vector is successfully constructed. The overexpression of miR-22 can inhibit the proliferation of HCT116 cells.
出处
《山西医科大学学报》
CAS
2015年第9期880-884,共5页
Journal of Shanxi Medical University
