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目的建立 TagMan 探针逆转录一实时荧光定量 PCR 技术测定喉癌组织中表皮生长因子受体(epidermal growth factor receptor,EGFR)mRNA 水平的方法,分析其应用价值。方法设计高特异性引物和探针,以 TagMan 探针逆转录-实时荧光定量 PCR 技术对32例喉鳞状细胞癌组织和癌旁组织学正常喉黏膜组织 EGFR mRNA 表达水平进行测定。构建含有 EGFR 的重组质粒,进行测序鉴定。结果经测序分析证实,含 EGFR 的重组质粒构建成功,喉癌组织中 EGFR mRNA 相对含量(中位数0.025)高于癌旁正常喉黏膜组织中 EGFR mRNA 相对含量(中位数0.008),两者间的差异经Wilcoxon 秩和检验,差异有统计学意义(P<0.01)。结论 TagMan 探针逆转录-实时荧光定量 PCR 技术可作为一种喉癌组织中 EGFR mRNA 水平定量测定的新方法。
Objective To establish a real-time quantitative reverse transcription-TagMan probe real-time fluorescence quantitative PCR method for the determination of epidermal growth factor receptor (EGFR) mRNA levels in laryngeal cancer method, and analyze its value. Methods High-specificity primers and probes were designed and the expression of EGFR mRNA in 32 specimens of laryngeal squamous cell carcinoma and normal laryngeal mucosa tissues was determined by TagMan probe reverse transcription-real-time fluorescence quantitative PCR. The recombinant plasmid containing EGFR was constructed and sequenced. Results Sequencing analysis confirmed that the recombinant plasmids containing EGFR were constructed successfully. The relative content of EGFR mRNA in laryngeal carcinoma (median 0.025) was higher than that in adjacent normal laryngeal mucosa (median 0.008) The differences between the two groups were statistically significant (P <0.01) by Wilcoxon rank sum test. Conclusion TagMan probe reverse transcription - real-time fluorescence quantitative PCR can be used as a new method for the quantitative determination of EGFR mRNA in laryngeal carcinoma.