HIV-1缺损慢病毒载体的高滴度制备及其介导的高效基因转移  

High-titer preparation of HIV-1-based defective lentivector and it mediated efficient gene transfer

作　　者：曾令兵[1,2,3] 张林[1] 孟彦[1] Yuanan Lu 叶林柏[3] 

机构地区：[1]中国水产科学研究院长江水产研究所,荆州434000 [2]Retrovirology Research Laboratory, Pacific Biosciences Research Center, University of Hawaii at Manoa, Honolulu, Hawaii 96822, USA [3]武汉大学生命科学学院病毒学国家重点实验室,武汉430072

出　　处：《微生物学报》2007年第6期1060-1065,共6页Acta Microbiologica Sinica

摘　　要：近来,慢病毒载体引起了极大关注,己成为转基因操作中重要的工具。用编码病毒组份的三质粒系统共转染293T包装细胞系,建立了大量制备HIV-1缺损慢病毒载体的方法,病毒载体的度可达到1.1×107IU/mL,离心浓缩可将载体滴度提高100倍以上。HIV-1缺损慢病毒载体可以高效转导人淋巴瘤等多种来源的细胞,RT-PCR检测显示外源基因GFP稳定表达达18个月以上,长期传代观测未检出p24抗原蛋白或可复制病毒。Lentivectors have drawn considerable attention recently and become important delivery vehicles for gene transfer manipulation. By Transiently co-transfecting 293T packaging cells with three DNA plasmids system encoding lentivector constituents, a protocol for bulky preparation of human immunodeficiency virus type-1 （HIV-1）-based defective lentivector with high titer has been established. Transient co-transfection of 293T packaging cells resulted in production of high-titer vector （1.1 × 10^7IU/ml）, which can be further concentrated over 100-fold through a single step centrifugation. The vector was capable of efficiently transducing a variety of cells from both primate and non-primate sources, including of human T-lymphoblastoid cell line. Long-term culture of vector transduced cells showed a stable expression of foreign gene over 18 months detected by RT-PCR. Assessment of potential generation of replication-competent virus revealed no detection of p24 antigen protein or infectious particles in vector-transduced cells.

关 键 词：HIV-1慢病毒载体 制备 高滴度 基因转移 效率 

分 类 号：Q78[生物学—分子生物学]