再组合克隆的文物挑战了对等离子体共变,选择和维护的既定信念
Courtney L Geer1,2,3, J Michael Charette1,2,3
1Department of Chemistry, Brandon University, Brandon, MB, Canada.
概括
网关克隆可以导致意想不到的共同转换. 这项研究确定了一种常见的"神秘"带作为埃舍里希亚大肠杆菌中的共转化Entry等离子体,挑战了标准的转化和选择信念.
科学领域:
- 分子生物学分子生物学
- 细菌遗传学 细菌遗传学
背景情况:
- 门户克隆是分子克隆的一个广泛使用的方法.
- 在Gateway克隆中的表达式克隆验证过程中经常观察到一个不明原因的带.
研究的目的:
- 为了确定在Gateway克隆过程中观察到的额外带的来源.
- 为了研究Escherichia coli中的等离子体共转化现象.
主要方法:
- 大肠杆菌与多个等离子体的转化.
- 在没有抗生素选择的情况下,分析细菌殖民地对等离子体的存在.
- 验证网关克隆结构的验证.
主要成果:
- "神秘"带是共变化的结果,其中多个等离子体被单个大肠杆菌细胞吸收.
- 在9%-29%的殖民地中,未被选择的pDONR输入等离子体与表达载体共转化.
- 尽管有等离子体不兼容性,但转化等离子体在大肠杆菌中保持不存在抗生素压力.
结论:
- 联合转换是Gateway克隆中一个重要的,以前未经记录的事件.
- 一种简单的选方法可以识别和消除共变剂.
- 这些发现需要重新评估细菌等离子体的转化,选择和维护原则.
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