在白狼中通过使用内源性促进体来适应基于CRISPR/Cas9的高效多重基因组编辑系统
Xiaoqi Zhu1, Weifeng Xu1, Bowen Liu1
1Joint International Research Laboratory of Water and Nutrient in Crop and College of Resource and Environment, Center for Plant Water-use and Nutrition Regulation and College of Life Sciences, Fujian Agriculture and Forestry University, Fuzhou, China.
Physiologia plantarum
|August 24, 2023
概括
这项研究成功地适应了白色狼 (Lupinus albus L.) 的CRISPR/Cas9基因编辑系统,使得在这个重要的作物中实现了高效的分子分析和功能基因组学研究.
科学领域:
- 植物分子生物学 植物分子生物学
- 作物基因组学作物基因组学
- 基因编辑技术的技术
背景情况:
- 白色狼 (Lupinus albus L.) 是一种重要的作物,以其高利用效率而闻名.
- 功能有限的基因组和分子分析工具阻碍了白狼的研究.
- CRISPR/Cas9系统在许多作物中彻底改变了基因编辑,但其在白狼中的应用尚未得到充分探索.
研究的目的:
- 适应和验证一个基于CRISPR/Cas9的多重基因组编辑系统,用于白狼.
- 评估原生促进剂对驱动sgRNA和Cas9表达的疗效.
- 为了研究白狼中目标基因的突变频率和类型.
主要方法:
- 使用Agrobacterium rhizogenes介导的转变来引入CRISPR/Cas9系统.
- 原生U3/U6和无素 (UBQ) 促进剂被用来驱动sgRNA和Cas9.
- 在Lalb_Chr05g0223881基因 (假定三酶) 内的两个点被准进行突变分析.
主要成果:
- 在T1 (6.25%-35%) 和T2 (50%-92.31%) 目标部位的T0毛发根中观察到有效突变.
- 观察到的突变包括插入,删除,替代和复杂变异,同时检测到两个目标的同时突变.
- 对sgRNA的LaU6.6促销剂和Cas9的LaUBQ12促销剂的组合产生了同卵性/双性突变的最高频率 (38.46%).
结论:
- 这项研究成功地建立了白色狼的CRISPR/Cas9基因组编辑工具箱.
- 这些发现显著有利于白狼的基础研究和功能基因组学.
- 优化促进体使用和目标sgRNA序列提高了该物种的CRISPR/Cas9编辑效率.
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