基于Ty反转移素元素的多重集成工具包用于Saccharomyces cerevisiae
Song Gao1,2,3,4, Weizhu Zeng1,3,4, Dong Li1,3,4
1Engineering Research Center of Ministry of Education on Food Synthetic Biotechnology, Jiangnan University, 1800 Lihu Road, Wuxi, Jiangsu, 214122, China.
Synthetic and systems biotechnology
|May 19, 2025
概括
这项研究引入了一种新的工具包,用于使用Ty元素集成在Saccharomyces cerevisiae (S. cerevisiae) 中高水平蛋白质的生产. 这种方法可以实现稳定,多副本的基因表达,从而提高蛋白质和代谢物产量.
科学领域:
- 合成生物学 合成生物学
- 分子生物学分子生物学
- 生物技术是生物技术.
背景情况:
- 在Saccharomyces cerevisiae (S. cerevisiae) 中实现高水平的蛋白质和代谢物生产需要稳定,高效的基因表达.
- 特定基因的基因组集成为持续的,高水平的蛋白质生产提供了一个强大的战略.
研究的目的:
- 通过将基因整合到Ty元素中来开发和验证S. cerevisiae中超高水平基因表达的工具包.
- 为了证明这个工具包在生产光蛋白和特定代谢物 (taxifolin) 的有效性.
主要方法:
- 利用S. cerevisiae CEN.PK2-1D中的五个Ty元素家族进行大规模的基因表达.
- 设计了九个选择性标记器,其中六个 (TRP1,LEU2,URA3,HIS5,natMX,hphMX) 允许在Ty站点稳定的高副本 (>15副本) 集成.
- 通过过度表达光蛋白和taxifolin生物合成途径基因来验证工具包.
主要成果:
- 获得了phiYFP的1.6g/L (268.1mg/g DCW) 的蛋白质标位,光强度比异常过度表达高3.3倍.
- 通过使用三个选择性标记物,成功将14个用于taxifolin生物合成的基因集成到三个Ty位点.
- 从葡萄糖中积累了277.6 mg/L的出租车folin.
结论:
- 开发的工具包可以使基因稳定,高复制地集成到Ty站点中,以提高S. cerevisiae中的蛋白质和代谢物生产.
- 这种方法与传统的插曲方法相比,显著提高了产量.
- 该工具包对蛋白质过度表达和复杂的代谢途径工程都有效.
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