Xu Meijuan, Liu Pingping, Chen Jiamin, Peng Anqi, Yang Taowei, Zhang Xian, Xu Zhenghong, Rao Zhiming
Key Laboratory of Industrial Biotechnology, Ministry of Education, School of Biotechnology, Jiangnan University, Wuxi, China.
Jiangnan University (Rugao) Food Biotechnology Research Institute, Rugao, China.
Front Bioeng Biotechnol. 2020 Mar 13;8:175. doi: 10.3389/fbioe.2020.00175. eCollection 2020.
The high yield mutants require a high-throughput screening method to obtain them quickly. Here, we developed an L-arginine biosensor (ARG-Select) to obtain increased L-arginine producers among a large number of mutant strains. This biosensor was constructed by ArgR protein and promoter, and could provide the strain with the output of bacterial growth via the reporter gene ; strains with high L-arginine production could survive in 10% sucrose screening. To extend the screening limitation of 10% sucrose, the sensitivity of ArgR protein to L-arginine was decreased. SYPA5-5 and its systems pathway engineered strain Cc6 were chosen as the original strains. This biosensor was employed, and L-arginine hyperproducing mutants were screened. Finally, the HArg1 and DArg36 mutants of SYPA5-5 and Cc6 could produce 56.7 and 95.5 g L of L-arginine, respectively, which represent increases of 35.0 and 13.5%. These results demonstrate that the transcription factor-based biosensor could be applied in high yield strains selection as an effective high-throughput screening method.
高产突变体需要一种高通量筛选方法来快速获得。在此，我们开发了一种L-精氨酸生物传感器（ARG-Select），以便在大量突变菌株中获得L-精氨酸产量增加的菌株。这种生物传感器由精氨酸阻遏蛋白（ArgR）和启动子构建而成，并能通过报告基因使菌株产生细菌生长的输出信号；L-精氨酸高产菌株能够在10%蔗糖筛选条件下存活。为了扩大10%蔗糖的筛选限度，降低了ArgR蛋白对L-精氨酸的敏感性。选择SYPA5-5及其系统途径工程菌株Cc6作为原始菌株。使用这种生物传感器筛选出了L-精氨酸高产突变体。最后，SYPA5-5和Cc6的HArg1和DArg36突变体分别能够产生56.7 g/L和95.5 g/L的L-精氨酸，产量分别提高了35.0%和13.5%。这些结果表明，基于转录因子的生物传感器可作为一种有效的高通量筛选方法应用于高产菌株的筛选。
