【摘要】 目的将唾液富组蛋白5(histatin5)的cDNA克隆至T载体,获取大量酶切后的目的片段。方法选用大肠杆菌偏爱密码子编码histatin 5氨基酸,设计5′端带酶切位点、3′端互补的两条引物,通过overlap PCR获取histatin 5的cDNA,将cDNA与T载体连接后转化大肠杆菌,筛选阳性克隆,PCR、酶切、测序鉴定。酶切重组T载体,获取目的片段。结果Histatin 5的cDNA正确克隆至T载体,无突变。结论克隆载体构建成功,有利于目的片段的酶切和回收。更多还原

【Abstract】 Objective To construct a cloning vector containing the cDNA of salivary histatin 5,acquire large amount of digested cDNA.Methods Histatin 5 was encoded by opitimal codon of E.coli,a pair of complementary primers were designed and used to synthsize the cDNA of histatin 5 by overlap PCR,the cDNA was flanked with EcoRⅠand SalⅠ sites.The cDNA was linked with T vector and transfered to E.coli,the positive recombinants were screened and identified by PCR,digestion and sequencing.Digested cDNA was obtained by digesting the positive recombinants.Results The cDNA of histatin 5 was correctly inserted into T vector wihout mutation.Conclusion Cloning vector was successfully constructed,digested cDNA could be harvested more easily.更多还原