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通过引入新突变来生成血友病A的小鼠模型,使用CRISPR/Nickase基因向系统

Mehdi Shamsara¹, Abbas Jamshidizad¹, Aidin Rahim-Tayefeh¹

¹Department of Animal Biotechnology, Institute Agricultural Biotechnology, National Institute of Genetic Engineering and Biotechnology, Tehran, Iran.

|September 18, 2023

概括

研究人员通过使用CRISPR-Cas9技术敲除凝血因子VIII (FVIII) 基因,开发出针对血友病A的新小鼠模型. 这些FVIII淘汰赛小鼠表现出疾病症状,适合进行体内治疗研究.

科学领域:

遗传学遗传学是一个
分子生物学分子生物学
血液学血液学血液学

背景情况:

血友病A是一种遗传性出血障碍,由缺乏凝血因子VIII (FVIII) 引起.
开发精确的动物模型对于研究血友病A机制和测试疗法至关重要.
现有的模型可能有局限性,需要创建新的,具有良好的特征的模型.

研究的目的:

通过在NMRI小鼠中敲除凝血因子VIII (FVIII) 基因来生成血友病A的小鼠模型.
使用CRISPR/Cas9 (D10A/nickase) 系统进行精确的基因编辑.
建立和验证表现出血友病A表型的遗传小鼠系.

主要方法:

在CRISPR/Cas9 (D10A/nickase) 系统中,有两个单向导RNA针对小鼠FVIII (mFVIII) 基因.
微注射CRISPR构造物到小鼠囊中,然后进行胚胎移植.
基因型定型,基因组测序和mFVIII活动测试用于识别和表征淘汰赛小鼠.
创始老鼠的杂交,以建立稳定的血友病A线.

主要成果:

成功生成了两个不同的FVIII淘汰赛 (FVIII_KO) 鼠标.
基因组分析证实了mFVIII基因中的基因破坏 (22-bp删除和23-bp插入).
FVIII_KO小鼠表现出血友病A的临床特征,包括过度和自发出血.
在FVIII_KO小鼠中观察到显著减少的mFVIII凝血活性.

结论:

克里斯普尔/尼卡斯系统有效地产生了可行的血友病A小鼠模型.
开发出的FVIII_KO小鼠线条准确地重复了疾病表型,并且是遗传的.
这些新型小鼠模型适用于血友病A疗法的临床前体内研究.

关键词:

克里斯普尔-Cas9是什么意思第八个因子血友病A型血友病A型血友病淘汰赛基因淘汰赛基因鼠标模型鼠标模型

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CRISPR

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