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Identification of differentially expressed genes in human dermal fibroblasts treated with sphingosylphosphocholine by suppression subtractive hybridization
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- Author:
- No author available
- Journal Title:
- CHINESE JOURNAL OF DERMATOLOGY
- Issue:
- 7
- DOI:
- 10.3760/j.issn:0412-4030.2006.07.004
- Key Word:
- 成纤维细胞;基因文库;神经鞘氨醇磷酸胆碱;抑制性消减杂交
Abstract： 目的利用抑制性消减杂交(SSH)技术构建神经鞘氨醇磷酸胆碱(SPC)对人真皮成纤维细胞的差异表达cDNA消减杂交文库,从中克隆出差异表达基因.方法原代培养人皮肤成纤维细胞;提取经SPC处理和未处理的成纤维细胞总RNA,合成cDNA,进行2次消减杂交及2次抑制性PCR,将产物与T/A载体连接,构建SPC作用于人真皮成纤维细胞的差异表达cDNA消减杂交文库,用RNA印迹验证阳性克隆,测序并登陆基因库寻找同源性基因.结果成功构建了SPC作用于人真皮成纤维细胞的差异表达cDNA文库;从实验组筛查的28个差异表达克隆中确定了6个差异表达基因,其中已知基因5个、新基因1个,分别为基质金属蛋白酶2,血小板反应蛋白1,催产素受体,锌指转录因子,铁传递蛋白受体等.结论神经鞘氨醇磷酸胆碱可上调人真皮成纤维细胞基质金属蛋白酶、血小板反应蛋白1、催产素受体、锌指转录因子、铁传递蛋白受体等基因表达.
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