中国当代医药Issue(32)：9-12,15,5.
USP22 ShRNA慢病毒载体的构建及鉴定
Construction and identification of ShRNA lentivirus vector in gene USP22
摘要
Abstract
Objective To construct and identify ShRNA lentivirus vector in gene USP22,and lay the foundation for fur-ther study of gene USP22 on mechanism of nasopharyngeal carcinoma. Methods Two special interference sequences targeting on coded sequence of gene USP22 were designed and synthesized.HpaⅠand XhoⅠas two sites of restriction enzyme were included at both ends of the sequence.Oligodeoxynucleotides was annealed to generate double strands of oligodeoxynucleotides.After 5’end phosphorylation,double strands of oligodeoxynucleotides containing restriction enzyme cutting site was cloned to pLentiLox3.7 (PLL3.7) lentiviral expression vector.The connection product was transformed and cultivated in order to extract plasmid.The extracted plasmid was identified by restriction enzyme digestion and electrophoresis,and sequencing was carried on only for correct plasmid after identification.The successfully-constructed lentiviral expression vector of pLL-USP22-shRNA and packaging plasmid were mixed and cotransfected into 293T cell. Conditions of green fluorescent protein (GFP) were observed under fluorescence microscope to determine virus titer and efficiency of infection. Results The sequencing showed that lentiviral expression vector of pLL-USP22-shRNA was successfully constructed.The lentiviral titer was 4í107 TU/ml after cotransfecting into 293T cell with packaging plasmid. Conclusion The lentivirus vector of USP22-ShRNA is successfully constructed by related techniques in the experi-ment,which lays the foundation for further study of biological function of gene USP22.关键词
USP22/慢病毒载体/构建/鉴定Key words
USP22/Lentiviral vector/Construction/Identification分类
医药卫生引用本文复制引用
余宏伟,廖志伟,庄雅靖,喻芳,周同冲..USP22 ShRNA慢病毒载体的构建及鉴定[J].中国当代医药,2014,(32):9-12,15,5.基金项目
广州医科大学附属肿瘤医院重大专项（2011-yz-06）；广州医科大学青年项目 （）
