The earthworm Metaphire posthuma were used as a model to assess the toxic potential of cadmium incorporated into the soil by environmental or human activities. The retention period of neutral red in the lysosomes ...The earthworm Metaphire posthuma were used as a model to assess the toxic potential of cadmium incorporated into the soil by environmental or human activities. The retention period of neutral red in the lysosomes of the coelomocytes was used as a biomarker. The viability of harvested coelomocytes by a non_invasive extrusion protocol was 93% with no alteration by the dye during experimentation. The control cells retained dye for 119 and 121 min in normal soil and KCl, respectively, whereas a linear decline in the retention time in the treated earthworm coelomocytes was observed. This illustrated that the presence of cadmium caused damage to the lysosomes of the coelomocytes.展开更多
目的建立SYBR Green Ⅰ实时荧光聚合酶链反应(PCR)检测survivin甲基化的方法。方法25例胃癌组织标本及与其配对的正常胃组织经甲基化敏感性限制性内切酶HpaⅡ和MspⅠ处理后,再用SYBR Green Ⅰ实时荧光PCR对survivin外显子1进行检测。实...目的建立SYBR Green Ⅰ实时荧光聚合酶链反应(PCR)检测survivin甲基化的方法。方法25例胃癌组织标本及与其配对的正常胃组织经甲基化敏感性限制性内切酶HpaⅡ和MspⅠ处理后,再用SYBR Green Ⅰ实时荧光PCR对survivin外显子1进行检测。实时荧光定量PCR检测survivin基因内含子2,用于监测经限制性酶消化的基因组DNA浓度变化,10倍系列稀释的基因组标准物用于检验实时PCR的敏感度,PCR产物通过凝胶电泳分析证实。结果经HpaⅡ酶切后,甲基化的目的基因PCR产物在熔解曲线上有Tm值为(91.5±0.5)℃的峰,电泳证实为338bp的条带。所有经酶消化的样本都能扩增出以Tm值(79.5±0.5)℃为特征的对照基因(survivin基因内含子2),表明基因组DNA未产生严重的非特异性降解。SYBR Green Ⅰ实时荧光PCR检测甲基化目的基因的灵敏度为100拷贝/μL。用以上新建的体系检测25例胃癌标本,发现其survivin基因外显子1的去甲基化频率为96%。结论SYBR Green Ⅰ荧光PCR法具有快速、准确、敏感、实时、简单和敏感的特点,是检测survivin外显子1甲基化状态的一种可靠的新方法。展开更多
文摘The earthworm Metaphire posthuma were used as a model to assess the toxic potential of cadmium incorporated into the soil by environmental or human activities. The retention period of neutral red in the lysosomes of the coelomocytes was used as a biomarker. The viability of harvested coelomocytes by a non_invasive extrusion protocol was 93% with no alteration by the dye during experimentation. The control cells retained dye for 119 and 121 min in normal soil and KCl, respectively, whereas a linear decline in the retention time in the treated earthworm coelomocytes was observed. This illustrated that the presence of cadmium caused damage to the lysosomes of the coelomocytes.
文摘目的建立SYBR Green Ⅰ实时荧光聚合酶链反应(PCR)检测survivin甲基化的方法。方法25例胃癌组织标本及与其配对的正常胃组织经甲基化敏感性限制性内切酶HpaⅡ和MspⅠ处理后,再用SYBR Green Ⅰ实时荧光PCR对survivin外显子1进行检测。实时荧光定量PCR检测survivin基因内含子2,用于监测经限制性酶消化的基因组DNA浓度变化,10倍系列稀释的基因组标准物用于检验实时PCR的敏感度,PCR产物通过凝胶电泳分析证实。结果经HpaⅡ酶切后,甲基化的目的基因PCR产物在熔解曲线上有Tm值为(91.5±0.5)℃的峰,电泳证实为338bp的条带。所有经酶消化的样本都能扩增出以Tm值(79.5±0.5)℃为特征的对照基因(survivin基因内含子2),表明基因组DNA未产生严重的非特异性降解。SYBR Green Ⅰ实时荧光PCR检测甲基化目的基因的灵敏度为100拷贝/μL。用以上新建的体系检测25例胃癌标本,发现其survivin基因外显子1的去甲基化频率为96%。结论SYBR Green Ⅰ荧光PCR法具有快速、准确、敏感、实时、简单和敏感的特点,是检测survivin外显子1甲基化状态的一种可靠的新方法。
