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  • 标题:Evolution of enzymes with new specificity by high-throughput screening using DmpR-based genetic circuits and multiple flow cytometry rounds
  • 本地全文:下载
  • 作者:Kil Koang Kwon ; Dae-Hee Lee ; Su Jin Kim
  • 期刊名称:Scientific Reports
  • 电子版ISSN:2045-2322
  • 出版年度:2018
  • 卷号:8
  • 期号:1
  • 页码:2659
  • DOI:10.1038/s41598-018-20943-8
  • 语种:English
  • 出版社:Springer Nature
  • 摘要:) were mutated randomly to construct a large diversity of specificity variations. Collection of candidate positives by cell sorting using flow cytometry and subsequent shuffling of beneficial mutations identified a critical hit with four mutations (D137P, F304D, V394L, and I396R) in the TIL sequence. The variant displayed one-thirteenth the level of TPL activity, compared with native TPLs, and completely lost the original TIL activity. The findings demonstrate that hypersensitive, Tx-based biosensors could be useful critically to generate new activity from a related template, which would alleviate the current burden to high-throughput screening.
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