利用CRISPRCas9技术构建模拟微管乙酰化和去乙酰化的定点突变果蝇

来源 :湖北省细胞生物学学会2015年会员代表大会暨学术研讨会 | 被引量 : 0次 | 上传用户:bjiyguang
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微管是细胞质骨架的重要组成成分,在维持细胞形态,调控细胞功能中发挥着重要的作用.细胞内存在多种微管翻译后修饰方式,其中乙酰化修饰因其存在的广泛性及在调控与微管功能相关的生物学过程中的独特作用而日益受到广泛的关注.例如,稳定微管,可以有效的避免微管破坏引起的神经退行性疾病相关的表型;抑制HDAC6的活性能够增加微管的乙酰化,从而部分挽救tau蛋白的毒性等.为了进一步的研究微管乙酰化的生物学功能,本研究旨在获得模拟微管第40位氨基酸乙酰化和去乙酰化的果蝇品系.利用CRISPR/Cas9定点突变技术,将果蝇的-tublin 84B的第40位赖氨酸(K)突变成谷氨酰胺(Q)模拟微管的乙酰化状态,突变成精氨酸(R)模拟微管的去乙酰化状态.经过测序鉴定,我们得到了这两种品系的纯合果蝇,这是第一次在生物体水平上实现原位定点突变模拟微管的乙酰化和去乙酰化状态,为后续微管乙酰化相关的研究提供了模式材料.
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