人神经营养因子3基因的分子克隆、表达及其产物的生物活性鉴定

以正常中国人血淋巴细胞染色体DNA为模板，PCR扩增出神经营养因子3（NT3）编码基因。将所得基因片段重组于M13噬菌体载体，筛选得到含中国人NT3基因的克隆。采用单链末端终止法测出其全部的核苷酸序列，该序列与国外文献所报道的完全一臻。将NT3编码基因亚克隆于杆状病毒表达载体，以在大肠杆菌内转座后的重组Bacmid大分子DNA转染悬浮培养的昆虫细胞后，在培养上清中检测到大量成熟型的NT3，后者对人神经母细胞瘤SH－SY5Y－T3具有较强的促进神经突起生长的作用，其生物学效应可被抗人NT3多克隆抗体所阻断。

Molecular cloning and expression of neurotrophin-3 gene and characterization of its expressed product

With the chromosomal DNA of human blood lymphocytes as template,neurotrophin-3 coding gene was amplified by PCR and recombinated into phage vector M13mp18RF,which was sequenced by using single-stranded DNA terminal termination method. The sequence of the cloned gene is completely the same as that of the literature reported. The NT-3 coding gene was subcloned in a baculovirus expression vector. A large quantity of mature NT3 was detected in the culture medium after the insect cells in suspension culture were transfected by high molecular weight of DNA transposed in E. coli. on human neuroblastoma cell line SH-SY5Y-T3. The NT3 in expressed supernatant have marked biological effect which can be blocked by anti-human NT3 polyclonal antibody.