Cas9 mRNA与gRNA在乳腺癌细胞中编辑LGMN基因的作用研究  

Cas9 mRNA and Guide RNA for Editing LGMN Gene in Breast Cancer Cells

作　　者：王越 彭保卫 Wang Yue;Peng Baowei(Dali University,Dali 671000,China)

机构地区：[1]大理大学,云南大理671000

出　　处：《广东化工》2023年第24期55-59,共5页Guangdong Chemical Industry

摘　　要：Legumain(或天冬氨酸内肽酶/AEP)是一种溶酶体半胱氨酸内肽酶,与多种癌症的侵袭和迁移行为增加相关。本研究通过设计Legumain(或AEP)特异性g RNA,构建了含有双U6启动子的g RNA质粒;构建了含T7启动子的g RNA质粒,优化RNA体外转录纯化方法。通过转染乳腺癌细胞株MDA-MB-231,对LGMN基因成功进行了基因敲除,比较Cas9 m RNA和g RNAs与Cas9和g RNAs双质粒在MDA-MB-231细胞中的基因编辑效率。结果表明醋酸铵纯化RNA效果更佳,Cas9 m RNA和g RNAs的基因编辑效率比Cas9和g RNAs的双质粒更高。Legumain(or aspartate endopeptidase/AEP)is a lysosomal cysteine endopeptidase associated with increased invasive and migratory behavior in a variety of cancers,Legumain(or AEP)specific gRNAs were designed.Plasmid containing dual U6 promoter-gRNAs was constructed;Plasmid containing T7 promoter-gRNAs was constructed,RNA purification methods were screened to optimize RNA purification.LGMN gene was successfully knocked out by transfection of breast cancer cell line MDA-MB-231,and the gene editing efficiency of Cas9 mRNA and gRNAs was compared with that of Cas9 and gRNAs double plasmids in MDA-MB-231 cells.The results showed that ammonium acetate was more effective in purifying RNA,and the gene editing efficiency of Cas9 mRNA and gRNAs was higher than that of Cas9 and gRNAs double plasmids.

关 键 词：CRISPR/Cas9 基因编辑 Legumain/AEP IVT 

分 类 号：R96[医药卫生—药理学]