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OALib Journal期刊
ISSN: 2333-9721
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稳定表达hmate1及共表达hmate1与hoct1或hoct2细胞模型的构建
, PP. 842-847
Keywords: 细胞模型,人多药及毒素外排转运体1,人有机阳离子转运体1,人有机阳离子转运体2,共表达
Abstract:
为构建稳定表达人多药及毒素外排转运体1(hmate1)的转基因细胞模型,提取人肾总mrna,经逆转录pcr获得hmate1cdna,借助hindⅲ、kpnⅰ两个酶切位点与pcdna3.1(+)重组获得重组质粒。将pcdna3.1(+)-hmate1重组质粒转染至mdck、mdck-hoct1和mdck-hoct2细胞中,经潮霉素b抗性筛选后,以4',6-二脒基-2-苯基吲哚(dapi)和n-甲基-4-苯基吡啶(mpp+)的积聚实验筛选获得具有良好hmate1功能的单克隆。测定筛选获得的细胞中转运体mrna的表达量,并表征其对二甲双胍的积聚或对西咪替丁的转运能力。结果表明,本研究构建的mdck-hmate1、mdck-hoct1/hmate1、mdck-hoct2/hmate1细胞模型均高表达hmate1mrna,mdck-hmate1细胞对二甲双胍的积聚为转染空载体细胞的17.6倍;mdck-hoct1/hmate1和mdck-hoct2/hmate1细胞对西咪替丁的净外排率分别为17.5和3.65。因此,本研究成功构建了稳定表达hmate1及共表达hmate1与hoct1或hoct2的细胞模型,可用于hmate1及其与hoct1或hoct2共同参与的药物转运或药物-药物相互作用的体外研究。
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