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发明公开
CN106967716A 双gRNA、双gRNA文库、双gRNA载体文库及其制备方法和应用 无效 - 驳回

双gRNA、双gRNA文库、双gRNA载体文库及其制备方法和应用
摘要：
本发明公开了一种双gRNA、双gRNA文库、双gRNA载体文库及其制备方法和应用。该双gRNA由第一启动子、gRNA‑a、crRNA支架序列、第二启动子和gRNA‑b顺次连接而成。该双gRNA文库包括针对靶基因的多个双gRNA。该双gRNA载体文库即由该双gRNA文库与载体连接而成。本发明的双gRNA中，gRNA‑a和gRNA‑b分别由不同的启动子控制其独立表达，并分别与靶基因上的两个靶位点互补配对，这使得该双gRNA能够招募Cas9核酸酶在两个sgRNA作用位点对靶基因进行剪切，有效提高靶基因的敲除效率。
摘要（英）：

The invention discloses a double gRNA, a double gRNA library, and a double gRNA vector library, a preparation method and applications thereof. The double gRNA is formed by sequentially linking a first promoter, gRNA-a, a crRNA scaffold sequence, a second promoter, and gRNA-b. The double gRNA library comprises a plurality of double gRNAs for a target gene. The double gRNA vector library is formed by linking the double gRNA library and a vector. According to the present invention, in the double gRNA, the independent expressions of the gRNA-a and the gRNA-b are controlled respectively by different promoters, and the gRNA-a and the gRNA-b are respectively complemented and paired with two target sites on the target gene, such that the double gRNA can recruit the Cas9 nuclease to cleave the target gent at the two sgRNA action sites so as to effectively improve the knockout efficiency of the target gene.

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