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

表1 本研究所用引物

Table 1 Primer sequences used in this paper

引物名称 Primer name 序列 Sequence (5′-3′) Cas9 F GACGACGACGACAAGGCCATGATGGACAAGAAGTACTCCA Cas9 R TCGACGGAGCTCGCTAGCGTCGCCGCCGAGCTGGGAGAGG Cas9NG F ACGACGACGACAAGGCCATGGACAAGAAGTACTCCATCGGC Cas9NG R CTCGAGTGCGGCCGCAAGCTTGTCGCCGCCGAGCTGGCTCAG sgRNA scaffold F GTTTTAGAGCTAGAAATAGCAAGTTAAAATAAGGCTAGTCCGTTATCAACTTGAAAAAG sgRNA scaffold R AAAAAGCACCGACTCGGTGCCACTTTTTCAAGTTGATAACGGACTAGCCTTATTTTAA esgRNA scaffold F GTTTCAGAGCTATGCTGGAAACAGCATAGCAAGTTGAAATAAGGCTAGTCCGTTATC esgRNA scaffold R AAAAAAGCACCGACTCGGTGCCACTTTTTCAAGTTGATAACGGACTAGCCTTATTTC ZmWx F CATACTTCTCCGGACCATACGGTAA ZmWx R TCCCTGCTGGGGTCCCACTC T7 F AAGCTAATACGACTCACTATAGGAGGTTCAGCTCCGGGTAGTGTTTTAGAGCTAGAAA eT7 F AAGCTAATACGACTCACTATAGGAGGTTCAGCTCCGGGTAGTGTTTCAGAGCTATGCT Target1 T7 F AAGCTAATACGACTCACTATAGGTTCAGCTCCGGGTAGTCGGGTTTCAGAGCTATGCT Target2 T7 F AAGCTAATACGACTCACTATAGCTCCGGGTAGTCGGAGAAGGGTTTCAGAGCTATGCT Target3 T7 F AAGCTAATACGACTCACTATAGCTTCTCCGACTACCCGGAGCGTTTCAGAGCTATGCT Target4 T7 F AAGCTAATACGACTCACTATAGTTCGCCTTCTCCGACTACCCGTTTCAGAGCTATGCT Target5 T7 F AAGCTAATACGACTCACTATAGGCCTTCTCCGACTACCCGGAGTTTCAGAGCTATGCT Target6 T7 F AAGCTAATACGACTCACTATAGCAGCTCCGGGTAGTCGGAGAGTTTCAGAGCTATGCT T7 R AAAAAGCACCGACTCGGTGCCACTTTTT