Port Fillip, Bullock Simon L
Cell Biology Division, MRC Laboratory of Molecular Biology, Cambridge, UK.
Nat Methods. 2016 Oct;13(10):852-4. doi: 10.1038/nmeth.3972. Epub 2016 Sep 5.
We present tRNA-based vectors for producing multiple clustered regularly interspaced short palindromic repeats (CRISPR) single guide RNAs (sgRNAs) from a single RNA polymerase II or III transcript in Drosophila. The system, which is based on liberation of sgRNAs by processing flanking tRNAs, permits highly efficient multiplexing of Cas9-based mutagenesis. We also demonstrate that the tRNA-sgRNA system markedly increases the efficacy of conditional gene disruption by Cas9 and can promote editing by the recently discovered RNA-guided endonuclease Cpf1.
我们展示了基于tRNA的载体，用于在果蝇中从单个RNA聚合酶II或III转录本产生多个成簇规律间隔短回文重复序列（CRISPR）单向导RNA（sgRNA）。该系统基于通过加工侧翼tRNA来释放sgRNA，允许基于Cas9的诱变进行高效多重化。我们还证明，tRNA-sgRNA系统显著提高了Cas9介导的条件性基因破坏的效率，并且可以促进最近发现的RNA引导的核酸内切酶Cpf1的编辑作用。
