基于CRISPR/Cas9核糖核蛋白体DNA定点内切酶体外活性建立高效基因型分析技术

王南,祁显涛,刘昌林,谢传晓,朱金洁

Establishment of an efficient genotyping technique based on targeted DNA endonuclease in vitro activity of CRISPR/Cas9 ribonucleoprotein

Nan WANG,Xian-Tao QI,Chang-Lin LIU,Chuan-Xiao XIE,Jin-Jie ZHU

图2 sgRNA、esgRNA体外转录及基于esgRNA/Cas9-RNP和esgRNA/Cas9NG-RNP检测体系的建立 A: sgRNA二级结构图; B: esgRNA二级结构图; C: RNA的10% Urea-PAGE胶图; D: PCR/RNP酶切检测; CK: 未加esgRNA和Cas蛋白的ZmWx野生型PCR产物。

Fig. 2 In vitro transcription of sgRNA and esgRNA, as well as the establishment of cleavage assay via esgRNA/Cas9-RNP and esgRNA/Cas9NG-RNP A: Predicted secondary structure of sgRNA; B: Predicted secondary structure of esgRNA; C: 10% Urea-PAGE of sgRNA and esgRNA; D: PCR/RNP cleavage assay; CK: ZmWx wild PCR product without esgRNA and Cas protein.