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目的 构建恶性疟原虫 CTP基因的真核表达载体 ,以便进一步研究其功能。 方法 根据 Genbank已发表恶性疟原虫CTP基因序列 (序列号为 X0 84 0 4 1) ,自行设计并合成了一对引物 ,通过聚合酶链反应扩增出 CTP基因 ,经 Hind 和 Bam H 消化后定向克隆入测序载体 p UC19,构建重组质粒 p UC19- CTP。经双酶切、PCR扩增和序列测定 ,证实插入片段与已知 CTP编码序列完全相同。用 Hind 和 Bam H 消化 p UC19- CTP,将双酶切下的 CTP编码基因片段定向亚克隆入真核表达载体 pc DNA3。 结果经双酶切和 PCR扩增鉴定证实 CTP基因正向插入真核表达载体 pc DNA3中。 结论 真核表达质粒 pc DNA3- CTP构建成功
Objective To construct eukaryotic expression vector of CTP gene of Plasmodium falciparum in order to further study its function. Methods A pair of primers was designed and synthesized according to the sequence of CTP gene of Plasmodium falciparum (GenBank No. X0 84 0 4 1). The CTP gene was amplified by polymerase chain reaction and digested with Hind and Bam H Cloned into the sequencing vector pUC19 to construct the recombinant plasmid pUC19-CTP. After double enzyme digestion, PCR amplification and sequence analysis, it was confirmed that the inserted fragment was exactly the same as the known CTP coding sequence. PUC19-CTP was digested with Hind and Bam H, and the double-digested CTP-encoding gene fragment was subcloned into the eukaryotic expression vector pcDNA3. Results Double digestion and PCR amplification confirmed that the CTP gene was inserted into the eukaryotic expression vector pcDNA3. Conclusion The eukaryotic expression plasmid pcDNA3-CTP was successfully constructed