Guangdong Key Laboratory of Fermentation and Enzyme Engineering, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, 510006, P. R. China.
Guangdong research center of Industrial enzyme and Green manufacturing technology, School of Biology and Biological Engineering, South China University of Technology, Guangzhou, 510006, P. R. China.
Sci Rep. 2017 Aug 11;7(1):7916. doi: 10.1038/s41598-017-08352-9.
Gene manipulation is essential for metabolic engineering and synthetic biology, but the current general gene manipulation methods are not applicable to the non-model strain Corynebacterium glutamicum (C. glutamicum) ATCC14067, which is used for amino acid production. Here, we report an effective and sequential deletion method for C. glutamicum ATCC14067 using the exonuclease-recombinase pair RecE + RecT (RecET) for recombineering via a designed self-excisable linear double-strand DNA (dsDNA) cassette, which contains the Cre/loxP system, to accomplish markerless deletion. To the best of our knowledge, this is the first effective and simple strategy for recombination with markerless deletion in C. glutamicum ATCC14067. This strategy provides a simple markerless deletion strategy for C. glutamicum and builds a solid basis for producer construction.
基因操作对于代谢工程和合成生物学至关重要，但目前的一般基因操作方法不适用于用于生产氨基酸的非模式菌株谷氨酸棒杆菌（Corynebacterium glutamicum）ATCC14067。在这里，我们报告了一种使用外切核酸酶-重组酶对谷氨酸棒杆菌 ATCC14067 进行有效且连续缺失的方法，该方法使用 RecE + RecT（RecET）对通过设计的自我切除线性双链 DNA（dsDNA）盒进行重组，该盒包含 Cre/loxP 系统，以完成无标记缺失。据我们所知，这是谷氨酸棒杆菌 ATCC14067 中首次具有无标记缺失的有效且简单的重组策略。该策略为谷氨酸棒杆菌提供了一种简单的无标记缺失策略，并为生产菌的构建奠定了坚实的基础。
