Hambor J E, Hauer C A, Shu H K, Groger R K, Kaplan D R, Tykocinski M L
Institute of Pathology, Case Western Reserve University, Cleveland, OH 44106.
Proc Natl Acad Sci U S A. 1988 Jun;85(11):4010-4. doi: 10.1073/pnas.85.11.4010.
A methodology was developed for stable gene transfer into cloned nontransformed human T lymphocytes. Stable high-level gene expression was achieved in cloned human T cells by using a self-replicating Epstein-Barr virus (EBV) episomal replicon. A comparison of five eukaryotic promoters established that the Rous sarcoma virus 3' long terminal repeat (RSV 3' LTR) and the lymphopapilloma virus (LPV) 5' LTR are optimal for episome-based expression in T cells. Effective (greater than 95%), selective, and reversible anti-sense RNA-mediated gene inhibition of a model T-cell-associated molecule (CD8) was achieved in a cytotoxic human T-cell clone by using an EBV episome-based, RSV 3' LTR-driven expression system. The linking of anti-sense RNA mutagenesis and T-cell cloning technologies should contribute significantly to studies of human T-cell function.
已开发出一种将基因稳定导入克隆的未转化人T淋巴细胞的方法。通过使用自我复制的爱泼斯坦-巴尔病毒（EBV）附加型复制子，在克隆的人T细胞中实现了稳定的高水平基因表达。对五种真核启动子的比较表明，劳氏肉瘤病毒3'长末端重复序列（RSV 3' LTR）和淋巴乳头瘤病毒（LPV）5' LTR最适合在T细胞中基于附加型的表达。通过使用基于EBV附加型、RSV 3' LTR驱动的表达系统，在细胞毒性人T细胞克隆中实现了对模型T细胞相关分子（CD8）有效（大于95%）、选择性和可逆的反义RNA介导的基因抑制。反义RNA诱变与T细胞克隆技术的结合应会对人T细胞功能的研究做出重大贡献。
