脑源性神经营养因子受体trkB在NIH 3T3细胞上的表达

Expression of Brain-derived Neurotrophic Factor Receptor trkB on NIH 3T3 Cells

构建了克隆有大鼠脑源性神经营养因子 (BDNF)受体 trk B全长基因的真核表达载体 pc DNA3 .1(+) -rat trk B.用脂质体介导法将重组载体转入小鼠 NIH3 T3细胞 ,在 m RNA和蛋白质水平检测到了 trk B基因在用 G41 8筛选到的抗性 NIH3 T3细胞中的表达 ,表达的 trk B蛋白定位于细胞膜上 .BDNF能够剂量依赖性地促进 NIH3 T3 -trk B细胞的增殖 ,说明表达的 trk B是有功能的 .该表达 trk B的 NIH3 T3细胞为研究BDNF的生理功能、活性测定和从噬菌体展示肽库中筛选

Recombinant eukaryotic expressing vector pcDNA3.1(+) rat trkB, constructed by cloning full length rat trkB cDNA into pcDNA3.1(+), was used to transfect murine NIH 3T3 cells by liposome mediated gene transfer method. The expressions of trkB at mRNA and protein levels were found in resistant NIH 3T3 cells selected by G418 with RT PCR and immunocytochemistry methods respectively. Moreover, the immunocytochemistry staining showed that trkB protein was expressed mainly on cytomembrane. BDNF could stimulate the proliferation of NIH 3T3 cells expressing trkB (NIH 3T3 trkB) in the dose dependent manner, suggesting that the expressed trkB possessed biological activity. NIH 3T3 trkB cells established in this work could be used as a cellular model for studying physiological functions, measuring activity of BDNF and selecting small BDNF mimicing peptides from phage displayed peptide libraries.