%0 Journal Article
%T The sectionalized DNA shuffling: an effective tool for molecular directed evolution of Meiothermus ruber TreS<br>分段DNA shuffling: 一种大分子海藻糖合酶有效的定向进化方法
%A Liu Yan-Chao
%A Wang Yu-Fan
%A Qian Ke-Fan
%A Zhang Jun
%A Xiao Chen-Peng
%A Xing Lai-Jun
%A Li Ming-Chun
%A <br>刘艳超
%A 王宇凡
%A 钱柯帆
%A 张峻
%A 肖辰鹏
%A 邢来君
%A 李明春
%J 微生物学通报
%D 2013
%I 
%X Objective] The gene M-treS from Meiothermus ruber CBS-01 encodes a trehalose synthase of 962 amino acids, named M-TreS. To improve its catalytic activity, we constructed a method of molecular directed evolution, the sectionalized DNA shuffling. Methods] Through two PCR steps with two pairs of partially complementary primers, the M-treS gene was parted into two sections. After the two sections shuffled respectively, a whole gene was obtained through the complementarity of the primers. This method was more feasible, with higher mutability than normal DNA shuffling. Results] Mutants were obtained after one round of the sectionalized DNA shuffling, in combination with error-prone PCR. The best mutant enzyme contained 6 amino acid substitutions, whose catalytic activity and efficiency were 1.6-fold and 2-fold of that of the wild type, respectively. In the 6 amino acid substitutions, 5 were caused by homologous recombination, and one by error-prone PCR. Conclusion] This study indicates that the sectionalized DNA shuffling is an effective tool for molecular directed evolution of macromolecular proteins.
%K Meiothermus ruber
%K TreS
%K Molecular directed evolution
%K Error-prone PCR
%K Sectionalized DNA shuffling<br>红色亚栖热菌
%K 海藻糖合酶
%K 定向进化
%K 易错PCR
%K 分段DNA
%K shuffling
%U http://www.alljournals.cn/get_abstract_url.aspx?pcid=90BA3D13E7F3BC869AC96FB3DA594E3FE34FBF7B8BC0E591&jid=78024727B5F4EF6AA9CA8E605B5FC464&aid=9C65539B20E123FC4A892042F01E2673&yid=FF7AA908D58E97FA&vid=1371F55DA51B6E64&iid=0B39A22176CE99FB&sid=FA3423FC1AE95C4E&eid=965F4E89CD0AFC30&journal_id=0253-2654&journal_name=微生物学通报&referenced_num=0&reference_num=0