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用于确定逆转录病毒滴度的颜色报告基因与耐药基因的系统比较。

Systematic comparison of a color reporter gene and drug resistance genes for the determination of retroviral titers.

作者信息

Sapp C M, Li T, Zhang J

机构信息

Department of Microbiology and Immunology, Markey Cancer Center, University of Kentucky, Lexington, KY 40536-0096, USA.

出版信息

J Biomed Sci. 1999 Sep-Oct;6(5):342-8. doi: 10.1007/BF02253523.

Abstract

Retroviral vectors usually contain drug resistance genes, which are used to select for infected cells and to determine the viral titers. The viral titer is referred to as colony-forming units (CFUs). Color reporter genes, such as the lacZ gene and the green fluorescent protein gene (gfp), have been widely used as markers in retroviral vectors. In this report, a simple and rapid method for the determination of retroviral titers has been developed. The number of viral particles capable of forming individual green cells per unit volume is defined as marker-forming units (MFUs). The MFUs determined by using gfp as a marker were found to be proportional to the CFUs obtained by using drug selection for five different drug resistance genes. In addition, after adjusting the time factor, the MFUs are higher than CFUs in viruses released from 30 stable helper cell lines. The lower titers determined by CFUs are likely due to the toxicity on transduced cells.

摘要

逆转录病毒载体通常包含耐药基因,这些基因用于筛选感染细胞并确定病毒滴度。病毒滴度称为集落形成单位(CFU)。颜色报告基因,如lacZ基因和绿色荧光蛋白基因(gfp),已被广泛用作逆转录病毒载体中的标记物。在本报告中,开发了一种简单快速的逆转录病毒滴度测定方法。每单位体积能够形成单个绿色细胞的病毒颗粒数量定义为标记物形成单位(MFU)。发现以gfp作为标记物测定的MFU与使用五种不同耐药基因进行药物筛选获得的CFU成正比。此外,调整时间因素后,从30个稳定辅助细胞系释放的病毒中,MFU高于CFU。CFU测定的较低滴度可能是由于对转导细胞的毒性。

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