摘要
目的建立分子信标定量检测Survivin基因mRNA表达的方法,对方法进行评估,并应用于临床。方法设计Survivin基因特异性分子信标探针和引物,以cDNA克隆重组质粒为标准品,建立Survivin基因分子信标定量检测方法;用该方法检测结直肠癌样本中Survivin基因mR- NA的表达量,分析mRNA表达量与临床资料之间的关系。结果所建立的分子信标定量检测方法,线性范围为1×103～1×1010拷贝数,批间变异为28．16％,批内变异为13．34％,灵敏度为42个拷贝数,平均回收率为109．83％;肠癌组织、癌旁组织和正常组织以及肠良性病变组织间比较,mR- NA表达量差异有统计学意义(P<0．05),肠癌样本中mRNA表达量与病理分型和淋巴结转移有关(P<0．05)。结论成功建立Survivin基因分子信标定量检测方法,为基因诊断提供新的定量检测方法。
Objective To develop and evaluate a real time fluorescent quantitative method for quantification of mRNA expression of survivin gene based on molecular beaoon technique. Methods A molecular beaoon probe and primer were designed and applied in the detection of survivin gene, while recombination plasmid oontaining the sequence of survivin was standard. The quantification of mRNA expression was detected in colorectal carcinoma, and the relationship between mRNA expression level and clinical data was analyzed. Results A linear standard curve was obtained between 1 × 10^3 and 1 × 10^10 copies. The inter and intra-assay coefficient variation （CV） were 28.16 % and 13.34 % respectively. The sensitivity of this assay was 42 copies. The average recovery was 109.83%. The quantification of mRNA expression of survivin was significantly higher in oolorectal carcinoma than in the other groups （P 〈 0.05）. There was a relationship between survivin gene expression with different pathological types and lymph node metastases. Conclusion The method can detect absolute quantification of mRNA expression of survivin gene, and can be applied to clinical diagnosis.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2006年第5期602-604,共3页
Chinese Journal of Experimental Surgery
基金
教育部重点资助项目(地方01006)
天津市卫生局基金资助项目(01KY31)
关键词
分子信标
绝对定量
SURVIVIN
MRNA表达
肠癌
Molecular beacon
Absolute quantification
Survivin gene
mRNA expression
Colorectal carcinoma
