酪氨酸羟化酶基因载体－pCMVTH　质粒的构建及在大鼠骨骼肌内的表达

RECONSTRUCTION OF TYROSINE HYDROXYLASE GENE VECTOR-pCMVTH AND EXPRESSION IN RAT MUSCLE CELLS

为用转基因方法治疗巴金森氏病大鼠模型，本研究采用分子克隆技术，将合成多巴胺的关键酶－酪氨酸羟化酶（ＴＨ）的基因，克隆进入以巨细胞病毒ＣＭＶ为启动子的载体质粒内，经限制性内切酶定位分析证实该重组的ＤＮＡ质粒的可靠性。携带ＴＨ基因的ＰＣＭＶＴＨ质粒以ＬＩＰＯ－ＦＥＣＴＩＮ介导，在培养的原代骨骼肌细胞中高效表达。本研究为进一步用转基因的细胞植入脑内以治疗巴金森氏病打下一定基础。

The purpose of this study was to determine whether the plasmid pCMVTH which reconstructed was able to show expression in rat muscle cells. We used tyrosine hydroxylasegene, which was from the plasmid of pKSTH as a transgene and cytomegalovirus (CMV),which was from the plasmid of pCMVLacZ as a cellular promoter to do subcloning. Theplasmid pCMVTH was identified by analysis of restriction endonuclease. Primary rat muscleCells were trans feeted with pCMVTH by lipofectin. About 30% myoblasts and myotubeshaving been cultured for 2 days showed expression of tyrosine hydroxylase gene wiht immunohistochmical staining. Our results may serve as a basis for in gene therapy inParkinson's disease by tranferring tyrosine hydroxylase gene into brain tissues.