蔡诚忠, 许洪卫, 谭龙益. 分时同步循环半定量RT—PCR检测胃癌细胞E—钙粘附素表达的方法学建立[J]. 中国肿瘤临床, 2006, 33(8): 424-426.
引用本文: 蔡诚忠, 许洪卫, 谭龙益. 分时同步循环半定量RT—PCR检测胃癌细胞E—钙粘附素表达的方法学建立[J]. 中国肿瘤临床, 2006, 33(8): 424-426.
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Cai Chengzhong, Xu Hongwei, Tan Longyi. The Establishment of A Methodology of Semiquantitative RT-PCR with Time-sharing Synchronized Circulation for Detection of E-cadherinLevels in Gastric Cancer Cells[J]. CHINESE JOURNAL OF CLINICAL ONCOLOGY, 2006, 33(8): 424-426.
Citation: Cai Chengzhong, Xu Hongwei, Tan Longyi. The Establishment of A Methodology of Semiquantitative RT-PCR with Time-sharing Synchronized Circulation for Detection of E-cadherinLevels in Gastric Cancer Cells[J]. CHINESE JOURNAL OF CLINICAL ONCOLOGY, 2006, 33(8): 424-426.
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分时同步循环半定量RT—PCR检测胃癌细胞E—钙粘附素表达的方法学建立

The Establishment of A Methodology of Semiquantitative RT-PCR with Time-sharing Synchronized Circulation for Detection of E-cadherinLevels in Gastric Cancer Cells

    摘要
  • 摘要: 目的:探索建立分时同步循环法分管半定量RT-PCR体系,并检测不同分化程度胃癌细胞中E-钙粘附素的相对表达量。方法:以胃癌细胞株HGC-27、MKN45、SCG-790为标本,抽提总RNA,合成第一链cDNA,优化PCR反应条件,建立分时同步循环法分管半定量RT-PCR反应体系,检测比较E-Cad的相对表达量。结果:采用LoTempPCR全息反应液优化,退火温度为85℃,循环数为33,分时同步循环数为8。检测得胃癌细胞的E-Cad表达量与分化程度成负相关,符合试验预期。结论:分时同步循环法分管半定量RT-PCR能克服同管竞争及平台期对试验的影响,理论上减少了试验误差,可以用以胃癌细胞E-Cad表达量的初步检测。

     

    Abstract: Objective: To establish a methodology of semiquantitative RT-PCR with time-sharing synchronized circulation in separated tube and to detect E-Cadherin levels in gastric cancer cells.Methods: Total RNA was extracted from gastric cancer lines HGC-27, MKN45 and SCG-790, and then the semiquantitatve RT-RCR with two-step-circle in separated tube to compare E-Cadherin levelof different malignant gastric cancer was conducted. Results: The most suitable annealing temperature was 85℃ with LoTemp PCR, totle circle was 33. Eight circles interval was found between two-steps.The expression of E-Cadherin detected by the method had the inverse relation with the malignant de-gree of gastric cancer cell, which accord with our anticipation. Conclusion: The method of Semiquanti-tatve RT-PCR with two-step circulation in a separated tube could decrease the error in the experiment theoretically. It could be used to detect the E- cadherin level of the gastric cancer cell.

     

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