设计和组装的117千克巴的Phaeodactylum tricornutum 光体基因组
Emma J L Walker1, Mark Pampuch1, Nelson Chang1
1Department of Biochemistry, Schulich School of Medicine and Dentistry, Western University, London, ON N6A 5C1, Canada.
Plant physiology
|December 19, 2023
概括
我们开发了高效的酵母组装方法,用于克隆Phaeodactylum tricornutum 光体基因组,从而实现合成生物学应用. 这些技术有助于光合作用真核生物的全基因组工程.
科学领域:
- 合成生物学 合成生物学
- 分子生物学分子生物学
- 基因组学就是基因组学.
背景情况:
- 合成生物学需要扩大可用的工程生物 (底盘) 的范围.
- 叶绿体基因组为工程光合作用核细胞提供了潜力,但缺乏高效的克隆和工程工具.
- 菲奥达克三角虫 (Phaeodactylum tricornutum) 化物基因组 (117-kb) 是此类工程工作的目标.
研究的目的:
- 开发和演示使用酵母组装的Phaeodactylum tricornutum хлоропласт基因组全基因组克隆的高效方法.
- 为合成生物学应用,使藻类叶绿体基因组的后续工程和操纵成为可能.
主要方法:
- 采用了两种酵母组装方法:从PCR碎片中溶解和从碎片中预克隆成等离子体.
- 在酵母 (Saccharomyces cerevisiae) 中的同源重组被用于组装重叠的叶绿体基因组片段和克隆载体.
- 克隆的 хлоропласт基因组是在大肠杆菌中传播的.
主要成果:
- 在最少的选 (10个酵母菌群) 中实现了90%至100%的克隆效率.
- 证明了在大肠杆菌中克隆的质细胞基因组的稳定维护和传播.
- 设计了一个克隆基因组与SapI站点进行线性化和整合设计者DNA磁带.
结论:
- 本文介绍的酵母组装方法为克隆大型质细胞基因组提供了有效的工具.
- 在大肠杆菌中稳定的传播促进了藻类质体的DNA传递机制的发展.
- 这些方法可以适应其他具有类似叶绿体基因组特征的物种.
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