文献类型：期刊文献

英文题名：Recycling selectable markers via Cre/loxP system for constructing Komagataella phaffii strains co-expressing multiple proteins

作者：Wang, Weixian Han, Minghai Zhu, Guofei Liu, Xiaohui Zhao, Tianming Ma, Xiaoyan Gong, Xun Xu, Cunbin

通信作者：Han, MH[1]|[144402eaf094eaf27b6d2]韩铭海;

机构：[1]Guizhou Inst Technol, Analyt & Testing Ctr, Guiyang 550000, Peoples R China;[2]Guizhou Inst Technol, Coll Food & Pharmaceut Engn, Guiyang 550000, Peoples R China

第一机构：贵州理工学院

通信机构：corresponding author), Guizhou Inst Technol, Coll Food & Pharmaceut Engn, Guiyang 550000, Peoples R China.|贵州理工学院食品药品制造工程学院;贵州理工学院;

年份：2024

外文期刊名：BIOTECHNOLOGY LETTERS

收录：;WOS:【SCI-EXPANDED(收录号:WOS:001172057700001)】；

基金：No Statement Available

语种：英文

外文关键词：Cre/loxP; Co-express multi-protein; Komagataella phaffii; Pichia pastoris; Selectable marker

摘要：Objective A convenient strategy was developed to recycle selectable markers using Cre/loxP system for constructing Komagataella phaffii strains co-expressing multiple proteins. Results A plasmid in this strategy was generated from pPICZ alpha A with integration of lox71-Sh ble-lox66. Firstly, the plasmid was inserted with one target protein gene and then transformed into K. phaffii KM71. Secondly, the auxiliary plasmid pPICZ alpha A/cre/his4 containing CRE recombinase gene was further chromosomally inserted to Sh ble gene therein. Finally, methanol induction was conducted to produce CRE for Cre/loxP-mediated recombination, and consequently, the sequence between lox71 and lox66 was deleted, leading to recycling of Zeo (R) and His- markers. Then the resulted strain expressing the one target protein was used as the host to which another target protein gene could be inserted by the same procedures. Conclusions With easy manipulation, the method was effective in recycling of the selectable markers, and consequently two protein genes were sequential integrated chromosomally and successfully co-expressed in the yeast.