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目的:探讨KOZAK序列对细胞周期调节蛋白1(CCNL1)基因在人乳腺癌细胞MDA-MB-231中表达的影响。方法:构建pcDNA3.1-CCNL1和pcDNA3.1-CCNL1-K(含有KOZAK序列)的真核重组表达质粒,采用Fugene HD转染试剂将重组表达质粒转染入MDA-MB-231细胞,并用荧光定量PCR和Western blot方法检测pcDNA3.1-CCNL1和pcDNA3.1-CCNL1-K重组质粒在MDA-MB-231细胞中表达的差异。结果:在MDA-MB-231细胞中质粒pcDNA3.1-CCNL1-K比空白对照细胞mRNA和蛋白质表达量要高;而质粒pcDNA3.1-CCNL1比空白对照细胞mRNA和蛋白质表达量也高。结论:在MDA-MB-231细胞中,质粒pcDNA3.1-CCNL1-K较pcDNA3.1-CCNL1的mRNA和蛋白表达水平均存在差异,KOZAK序列能够提高CCNL1基因在MDA-MB-231细胞中的表达。
Objective: To investigate the effect of KOZAK sequence on the expression of CCNL1 gene in human breast cancer cell line MDA-MB-231. Methods: Eukaryotic recombinant plasmids containing pcDNA3.1-CCNL1 and pcDNA3.1-CCNL1-K (containing KOZAK sequence) were constructed and transfected into MDA-MB-231 cells by Fugene HD transfection reagent. The difference of expression of pcDNA3.1-CCNL1 and pcDNA3.1-CCNL1-K recombinant plasmid in MDA-MB-231 cells was detected by quantitative PCR and Western blot. Results: The mRNA and protein expression levels of pcDNA3.1-CCNL1-K in MDA-MB-231 cells were higher than those in blank control cells. The expression of pcDNA3.1-CCNL1 in mRNA and protein was also higher than that in blank control cells. Conclusion: The mRNA and protein expression levels of pcDNA3.1-CCNL1-K in pcDNA3.1-CCNL1-K are different from those in pcDNA3.1-CCNL1 in MDA-MB-231 cells. KOZAK sequence can increase the expression of CCNL1 in MDA-MB-231 cells expression.