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利用pGEM-T载体对CppB基因PCR扩增产物进行直接克隆,除常规方法中的限制性内切酶可以节省外,发现该方法不仅过程简便,操作易行,而且可以用PCR方法筛选重组子,克隆的阳性率较常规方法高。DNA序列测定结果证明在PCR产物3′末端的确出现了一个不与模板互补的“A”末端,它可以与PGEM-T载体T结合。PCR方法筛选重组,克隆的阳性率较常规方法高。DNA序列测定结果证明在PCR产物克隆片段的基因序列与文献报道相一致,这为获得淋球菌隐蔽性质粒CppB基因PCR诊断标准阳性模板,了解CppB基因功能奠定了基础。
The direct cloning of the PCR amplification product of CppB gene by the pGEM-T vector can save the conventional restriction endonuclease and find that the method is not only simple and easy to operate, but also can be used to screen the recombinants by the PCR method, Clone positive rate higher than the conventional method. The results of the DNA sequencing proved that an “A” non-complementary template was indeed found at the 3 ’end of the PCR product, which can bind to the PGEM-T vector T. PCR screening and recombination, the positive rate of cloning than the conventional method. The results of DNA sequencing confirmed that the sequence of the cloned fragment in PCR product was consistent with that reported in the literature, which laid the foundation for understanding the CppB gene function by obtaining the diagnostic template of CppB gene in Neisseria gonorrhoeae.