Crispr/Cas9 慢病毒

 

服务简介

       

        Cas9蛋白的CDS长达4kb,克隆难度和包毒难度都相对大,将Cas9基因高效导入细胞是应用Cas9/CRISPR系统进行基因敲除的难点之一,极大地限制可供选择的将基因导入细胞的方法。并且在用CRISPR/Cas9系统进行基因敲除时,还需要同时转入识别靶点的gRNA,筛选阳性细胞的抗性基因或荧光基因,用同源重组法进行基因敲除,还需要导入同源重组模板。如此多的基因共同表达,很难得到较高的基因编辑效率,造成后续的阳性克隆筛选和检测工作难度大。

       汉恒生物独创性研发出慢病毒Cas9表达体系,该体系采用慢病毒体系先在细胞中稳定表达Cas9蛋白,构建稳定表达Cas9蛋白的细胞系,再在该细胞系的基础上导入gRNA和基因敲除实验中用到的其它原件用于特异性基因敲除。

 

 产品优势

 

a、实验方案设计更灵活。由于不必表达大蛋白Cas9,可以根据细胞特性选择化学转染、腺病毒、腺相关病毒等多种方法导入或敲除相关基因;

 

b、提高基因敲除效率。在稳定表达Cas9蛋白的细胞系上进行基因敲除比瞬时转染Cas9进行基因敲除的效率更高;

 

c、可对同一细胞进行多个基因敲除实验。对于需要在同一个细胞系中进行多个基因的编辑或需要长期用一个细胞模型进行多项基因研究,先构建一个Cas9蛋白稳定表达的细胞系可显著提高后续实验的效率,降低实验难度。

 

 

 

服务特点

 

a、适用于在同一个细胞系中需要进行多个基因敲除的实验;

b、基因敲除效率比直接质粒转染更高;

c、提供多种不同荧光和抗性标记的慢病毒表达载体,更易筛选到基因敲除成功细胞;

d、接受cas9蛋白稳定表达不同细胞系定制。

 

 

服务流程

 

 

 

 

A、cas9载体构建及慢病毒包装

     将cas9基因CDS区克隆至慢病毒载体,并进行cas9慢病毒包装。

B、稳定株筛选

     将cas9慢病毒感染特定的目的细胞,进行cas9稳定表达的细胞株筛选。

 

 

参考文献

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