在Komagataella phaffii中作为外来遗传物质存储库的超数合成染色体
Dariusz Abramczyk1, Maria Del Carmen Sanchez Olmos2, Adan Andres Ramirez Rojas2
1School of Chemistry, University of Edinburgh, Edinburgh, UK. dabramcz@ed.ac.uk.
Microbial cell factories
|December 16, 2023
概括
研究人员在Komagataella phaffii (Pichia pastoris) 酵母中设计了一种稳定的人工"纳米"染色体,以实现高效的生物制造. 这项技术可以通过促进多个基因的整合而产生复杂的人类蛋白质,如补充因子H,而不会导致基因组不稳定.
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:
- 科马加泰拉 (Pichia pastoris) 是一种用于高密度发酵和高效重组蛋白质分泌的甲基转化酵母.
- 在K. phaffii中,基因工程旨在增强具有成本效益的生物制造过程.
- 小而稳定的人工染色体可以实现多个基因的整合,而不会扰乱原生染色体或耗尽选择标记.
研究的目的:
- 开发和描述一个稳定的,线性的人工.
主要方法:
- 在K. phaffii中构建一个15-25kb线性纳米染色体,其中包括一个中心粒,自主复制序列和端粒.
- 损害非同类末端连接 (KU70-Knockout),以确保几代人的纳米染色体稳定性.
- 在纳米染色体的着陆区内开发了一种"英寸虫"序列基因整合系统,利用两个抗生素耐药性标记物.
主要成果:
- 整个基因组测序证实了纳米染色体 (每细胞1个副本) 在多代人间的持久性.
- 从纳米染色体成功表达蛋白二硫化异构酶 (PDI),并随后与小鼠和人类补充因子H (CFH) 共同表达PDI.
- 作为补充系统的调节者,人类分泌的CFH的功能已被证明.
结论:
- 现在K. phaffii可以从稳定,线性人造染色体上的基因中产生人类蛋白质.
- 纳米染色体作为多个工程基因的稳定存储库,使密集的K. phaffii修饰能够在不损害基因组完整性或菌株性能的情况下实现.
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