Stable gene transfer and expression in human primary T cells by the Sleeping Beauty transposon system
Stable gene transfer and expression in human primary T cells by the Sleeping Beauty transposon system
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DOI：
10.1182/blood-2005-05-2133
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发表时间：
2006-01-15
期刊：
影响因子：
20.3
通讯作者：
Zhou, XZ
中科院分区：
文献类型：
--
作者：
Huang, X;Wilber, AC;Zhou, XZ
睡美人（SB）转座子系统是一种非病毒DNA递送系统，其中转座酶指导SB转座子整合到基因组中的TA二核苷酸位点中。为了确定SB转座子系统是否可以介导人T细胞中的稳定基因表达，用携带DsRed报告基因的SB载体核转染原代外周血淋巴细胞（PBL）。在与SB转座子相同的分子上（顺式）或在与SB转座子分开的分子上（反式）含有SB转座酶的质粒介导人原代T细胞中长期和稳定的报告基因表达。转座子：染色体连接测序证实，稳定的基因表达是由于SB介导的转座。在其它研究中，使用SB转座子系统成功转染PBL，并显示稳定表达融合蛋白，所述融合蛋白由（1）用于阳性T细胞选择的表面受体和（2）用于在化疗后消除转染的T细胞的"自杀"基因组成。本研究首次证明SB转座子系统可以介导人原代PBL中的稳定基因转移，这可能有利于基于T细胞的基因治疗。
The Sleeping Beauty (SB)transposon system is a nonviral DNA delivery system in which a transposase directs integration of an SB transposon into TA-dinucleotide sites in the genome. To determine whether the SB transposon system can mediate stable gene expression in human T cells, primary peripheral blood lymphocytes (PBLs) were nucleofected with SB vectors carrying a DsRed reporter gene. Plasmids containing the SB transposase on the same molecule as (cis) or on a molecule separate from (trans)the SB transposon mediated long-term and stable reporter gene expression in human primary T cells. Sequencing of transposon:chromosome junctions confirmed that stable gene expression was due to SB-mediated transposition. In other studies, PBLs were successfully transfected using the SB transposon system and shown to stably express a fusion protein consisting of (1) a surface receptor useful for positive T-cell selection and (2) a '' suicide '' gene useful for elimination of transfected T cells after chemotherapy. This study is the first report demonstrating that the SB transposon system can mediate stable gene transfer in human primary PBLs, which may be advantageous for T-cell-based gene therapies.