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大规模的全基因组优化和预测Cre重组酶系统用于小鼠精确的基因组操纵

Vishnu Hosur¹, Valerie Erhardt¹, Elli Hartig¹

¹The Jackson Laboratory for Mammalian Genetics.

Research square

|July 16, 2024

概括

在小鼠中优化Cre-Lox重组需要仔细考虑诸如loxP位点距离和Cre-driver菌株选择等因素. 了解这些元素可以提高效率,并降低产生转基因小鼠模型的成本.

科学领域:

遗传学和基因组学在
分子生物学分子生物学
动物模型动物模型

背景情况:

在老鼠遗传学中,Cre-Lox重组系统对于条件基因操纵至关重要.
挑战包括高成本,时间投资和不可预测的重组事件.
需要进行系统分析,以优化Cre-Lox用于生成小鼠模型的效率.

研究的目的:

在小鼠中进行影响CRE媒介重组的因素的全基因组分析.
为了确定有效和可重复的Cre-Lox重组的最佳条件.
为生成条件淘汰赛小鼠和其他遗传模型提供指导.

主要方法:

在C57BL/6J背景上的ROSA26位点生成了11个具有条件等位基因的新型小鼠菌株.
系统地评估了loxP间距离,突变的loxP位点,雄性,染色体位置和育种者年龄对重组的影响.
利用Cre-driver菌株在各种条件下评估重组效率.

主要成果:

培养驱动菌株选择显著影响重组效率.
最佳的重组发生在距离1-4kb的loxP位点;突变的loxP位点在1-3kb之间是有效的.
在距离≥15kb (野生型loxP) 或≥7kb (突变型loxP) 时,完全重组是受阻的;育种年龄 (8-20周) 和异构基提高了效率.

结论:

在有效的Cre-Lox重组中,LoxP间的距离,LoxP位点类型,Zygosity和繁殖者年龄至关重要.
保持loxP间距离≤4kb可以确保有效的重组,无论染色体位置如何.
这项研究提供了优化Cre-Lox策略的重要数据,减少了生成小鼠模型的时间和成本.

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