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首页> 中文期刊> 《中国病理生理杂志》 >siRNA-survivin与GRIM-19共表达质粒的构建及其对前列腺癌DU145细胞生长的抑制作用

siRNA-survivin与GRIM-19共表达质粒的构建及其对前列腺癌DU145细胞生长的抑制作用

         
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摘要

ADM: To construct the recombinant plasmid that expresses siRNA — survivin and GRIM — 19 simultaneously , and to identify the validity of the recombinant plasmid and observe its effect on expression of survivin and GRIMrn- 19 and proliferation ability of prostate cancer DU145 cells. METHODS: The recombinant plasmid coexpressing siRNArn— survivin and GRIM — 19 was constructed using gene cloning technique. The prostatic cancer DU145 cells were transfect-ed with the coexpression plasmid and control plasmids. Survivin and GRIM — 19 mRNA expression was detected by semi — quantitative RT - PCR. The proliferation ability affected by coexpression plasmid was measured by MTT assay. RESULTS : The coexpression plasmid pGRIM — 19 — si — survivin was successfully constructed according to DNA recombinant technique and identified through restriction enzyme digestion and plasmid sequencing. Compared with the mock, survivin mRNA expression levels were 0. 55 ±0. 05,0. 62 ±0. 08 and 0. 35 ±0. 05 in psi - survivin, pGRIM - 19 and pGRIM - 19rn- si - survivin groups, respectively. Compared with psi - survivin and pGRIM - 19, pGRIM - 19 - si - survivin inhibited survivin mRNA expression markedly ( P < 0. 05 ), while the expression levels of GRIM — 19 mRNA were 1. 93 ±0. 14, 2. 57 ±0. 20 and 4. 12 ±0. 21 in psi - survivin, pGRIM - 19 and pGRIM - 19 - si - survivin groups, respectively ( P < 0. 01 ). Compared with pGRIM — 19 group, pGRIM — 19 — si — survivin enhanced GRIM — 19 mRNA expression more obviously ( P < 0. 05 ). After transfection for 48 h, the proliferation rates were 58. 0% ± 7. 2% , 62. 1 % ± 6. 1 % and 50. 2%rn± 4. 8% in the 3 experiment groups compared with the mock ( P < 0. 05 ). After transfection for 72 h, the proliferation rate were 43. 4% ±4. 3% , 51. 3% ±6. 7% and 26. 8% ±7. 1% in experiment groups compared with the mock ( P <0. 05 ). Compared with psi — survivin and pGRIM — 19, pGRIM — 19 — si — survivin significantly inhibited the cell growth ( P < 0. 05 ). CONCLUSION: Transfection of coexpression plasmid pGRIM — 19 — si — survivin dramatically changes the mRNA expression of survivin and GRIM - 19 and inhibits the cell growth.%目的:构建siRNA-survivin与GRIM-19共表达质粒pGRIM-19-si-survivin并鉴定,观察pGRIM-19-si-survivin质粒对人前列腺癌DU145细胞survivin和GRIM-19 mRNA表达水平及细胞增殖能力的影响.方法:根据前期工作,利用基因重组技术构建siRNA-survivin与GRIM-19共表达质粒,将成功构建的pGRIM-19-si-survivin质粒转染人前列腺癌DU145细胞株,应用半定量RT-PCR方法检测细胞内survivin和GRIM-19 mRNA表达水平,应用MTT法观察其对细胞增殖能力的影响.结果:(1)应用酶切及质粒测序法鉴定,证明成功构建siRNA-survivin与GRIM-19共表达质粒.(2)与mock组比较,psi-survivin、pGRIM-19及pGRIM-19-si-survivin组survivin mRNA表达水平分别为0.55±0.05、0.62±0.08和0.35±0.05,差异显著(P<0.01);与psi-survivin和pGRIM-19组比较,pGRIM-19-si-survivin的抑制survivin表达作用明显增强(P<0.05);psi-survivin、pGRIM-19和pGRIM-19-si-survivin组GRIM-19表达增强,分别为mock组的1.93±0.14、2.57±0.20 和4.12±0.21(P<0.01),与pGRIM-19组比较,pGRIM-19-si-survivin增强GRIM-19表达作用更明显(P<0.05).(3)转染48 h后,与mock组比较,psi-survivin、pGRIM-19和pGRIM-19-si-survivin 3组细胞增殖率分别为58.0%±7.2%、62.1%±6.1%和50.2%±4.8%,差异显著(P<0.05);转染72 h后,与mock组比较,3组细胞增殖率分别为43.4%±4.3%、51.3%±6.7%和26.8%±7.1%,差异明显(P<0.05,P<0.01);与psi-survivin和pGRIM-19组比较,pGRIM-19-si-survivin抑制作用明显增强(P<0.05).结论:成功构建siRNA-survivin和GRIM-19共表达质粒pGRIM-19-si-survivin;该质粒抑制survivin表达并增强GRIM-19表达,对前列腺癌DU145细胞具有协同增殖抑制效应.

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  • 来源
    《中国病理生理杂志》 |2012年第7期|1241-1246|共6页
  • 作者

    沈维高; 赵丽晶; 盖晓东; 芦丽莉; 葛贺; 陈立松; 刘艳波; 赵雪俭;

  • 作者单位

    北华大学基础医学院病理生理学教研室,吉林,吉林,132013;

    吉林大学前列腺疾病防治研究中心,吉林,长春,130021;

    北华大学基础医学院病理生理学教研室,吉林,吉林,132013;

    北华大学基础医学院病理生理学教研室,吉林,吉林,132013;

    北华大学基础医学院病理生理学教研室,吉林,吉林,132013;

    北华大学基础医学院病理生理学教研室,吉林,吉林,132013;

    北华大学基础医学院病理生理学教研室,吉林,吉林,132013;

    吉林大学前列腺疾病防治研究中心,吉林,长春,130021;

  • 原文格式 PDF
  • 正文语种 chi
  • 中图分类 前列腺肿瘤;
  • 关键词

    前列腺肿瘤; 存活蛋白; GRIM-19;

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