两种pUC18高效T载体的构建  

Construction of Two Highly Efficient pUC18 T-vectors

作　　者：刘勇[1] 王立良[2] 袁力勇[1] 陈尔佳[1] 庄俊英[1] 孙茂盛[1] 戴长柏[1] 

机构地区：[1]中国医学科学院中国协和医科大学医学生物学研究所,昆明650118 [2]中国医学科学院中国协和医科大学基础医学研究所,北京100005

出　　处：《中国生物化学与分子生物学报》2000年第4期562-564,共3页Chinese Journal of Biochemistry and Molecular Biology

摘　　要：Two T vectors were generated by restriction endonuclease Xcm Ⅰ instead of Taq or other DNA polymerases to create the 3’T over hangs.A fragment of adenoviral genome position 10659-11865 was amplified by PCR and Xcm Ⅰ recognition sites were introduced to both ends of the PCR product.This fragment was cloned into the Sma Ⅰ site of pUC18 or the linearized pUC18 with its polylinker deleted.The recombinant plamids were cleaved with Xcm Ⅰ.the larger fragments generated which had 3’T over hangs at both ends were used as T vctors.Genes of rotavirus VP7 and the plasminogen k5 were successfully cloned into these two T vectors with recombination efficiency (recombinants/transformants×100%) of 100%,no blue/white clolny screening assay was needed.Two T vectors were generated by restriction endonuclease Xcm Ⅰ instead of Taq or other DNA polymerases to create the 3'T over hangs.A fragment of adenoviral genome position 10659-11865 was amplified by PCR and Xcm Ⅰ recognition sites were introduced to both ends of the PCR product.This fragment was cloned into the Sma Ⅰ site of pUC18 or the linearized pUC18 with its polylinker deleted.The recombinant plamids were cleaved with Xcm Ⅰ.the larger fragments generated which had 3'T over hangs at both ends were used as T vctors.Genes of rotavirus VP7 and the plasminogen k5 were successfully cloned into these two T vectors with recombination efficiency (recombinants/transformants×100%) of 100%,no blue/white clolny screening assay was needed.

关 键 词：PUC18 T载体 PCR 基因克隆 XcmⅠ 

分 类 号：Q782[生物学—分子生物学]