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目的构建CD147siRNA表达载体,分析CD147在皮肤肿瘤浸润和转移中的作用机制。方法根据基因库上的CD147cDNA序列,设计并合成两端含有酶切位点的64个碱基的寡核苷酸链。寡核苷酸链退火后用T4DNA连接酶连接到线性化的pSUPER质粒中,并对重组质粒(命名为pSUPER/CD147siRNA)进行酶切、PCR及测序鉴定。结果PCR扩增片段与预期结果相符,双酶切证实CD147siRNA表达载体克隆构建成功,插入片段测序结果与合成的siRNA结果一致。结论成功构建CD147siRNA表达载体,CD147可为治疗肿瘤开辟新途径。
Objective To construct CD147 siRNA expression vector and analyze the mechanism of CD147 in the invasion and metastasis of cutaneous tumors. Methods According to the CD147 cDNA sequence on the gene bank, a 64 - base oligonucleotide chain containing restriction enzyme sites at both ends was designed and synthesized. The oligonucleotide strands were annealed and then ligated into the linearized pSUPER plasmid using T4 DNA ligase. The recombinant plasmid (named pSUPER / CD147 siRNA) was digested with restriction endonuclease, PCR and sequencing. Results The PCR amplified fragment was consistent with the expected result. Double enzyme digestion confirmed that the cloning of CD147 siRNA expression vector was successful. The results of insert sequencing were consistent with those of the synthesized siRNA. Conclusion The CD147siRNA expression vector was successfully constructed and CD147 could open up a new way for the treatment of tumors.