Integration efficiency of a hybrid adenoretroviral vector.
Integration efficiency of a hybrid adenoretroviral vector.
复制标题
混合腺逆转录病毒载体的整合效率。
DOI：
10.1016/s0006-291x(02)02801-2
复制
发表时间：
2003
影响因子：
3.1
通讯作者：
Baum,BruceJ
中科院分区：
文献类型：
--
作者：
Zheng,Changyu;Wang,Jianghua;Baum,BruceJ
杂交载体AdLtr-Luc介导基因组整合的频率[NAT.生物技术。18(2000)176-180]不详。为了解决这一问题，我们使用AdLTR-Luc骨架和增强的红色荧光蛋白cDNA构建了AdLTR-RED。动力学研究表明，AdLTR-RED和常规腺病毒载体AdCMV-RED进入和转导上皮细胞的情况相似。AdLTR-RED体外整合频率为8.0%(36/450；67个/细胞)。用AdCMV-red检测，整合阳性克隆为0/549个。成年大鼠股静脉注射AdLTR-Luc或AdCMV-Luc(1011粒)7天后，制备脾细胞，用刀豆蛋白A刺激后进行FISH检测。有丝分裂的大鼠脾细胞中有5-11%存在AdLTR-Luc整合，而AdCMV-Luc未见明显整合。这些数据为基因组与该杂交载体整合的频率提供了第一个定量证据。
The frequency with which the hybrid vector AdLTR-luc mediates genomic integration [Nat. Biotech. 18 (2000) 176–180] is unknown. To address this, we constructed AdLTR-red, using the AdLTR-luc backbone and the enhanced red fluorescence protein cDNA. Kinetic studies showed that AdLTR-red and a conventional adenoviral vector, AdCMV-red, entered and transduced epithelial cells comparably. AdLTR-red integration frequency in vitro, i.e., the percentage of red clones after 10–12 doubling times from limiting dilutions, was 8.0% (36/450; at 67particles/cell). With AdCMV-red, 0/549 clones were integration-positive. Seven days after AdLTR-luc or AdCMV-luc (1011particles) femoral vein administration to adult rats splenocytes were prepared, stimulated with concanavalin A, and examined by FISH. AdLTR-luc integration occurred in 5–11% of mitotic rat splenocytes, while no unequivocal integration was found with AdCMV-luc. These data provide the first quantitative evidence of the frequency for genomic integration with this hybrid vector.