Rb2/p130寡核苷酸基因芯片的制备及肺癌手术标本的检测  

Preparation of Rb2/p130 oligonucleotide gene chips and detection of Rb2/p130 oligonucleotide gene chips in surgical specimens of patients with lung cancer

作　　者：安耀武[1] 于海建[1] 李龙芸[2] 

机构地区：[1]南方医科大学附属花都医院,广东广州510800 [2]北京协和医院,北京100730

出　　处：《甘肃医药》2010年第6期601-605,共5页Gansu Medical Journal

摘　　要：目的：用基因芯片方法检测肺癌标本中Rb2/p130基因的点突变情况,为肺癌诊断探索快速、准确的基因检测途径。方法：收集肺癌患者的手术标本,提取基因组DNA,检索Rb2/p130基因的DNA序列后,设计并合成探针和引物序列,将探针按一定顺序点样于醛基化玻片,制备成基因芯片。用荧光标记引物对肺癌标本DNA进行PCR扩增,扩增产物与基因芯片杂交,通过对杂交信号的检测判断是否发生热点突变,并用测序方法对突变位点进行验证。结果：在30例用寡核苷酸基因芯片的方法检测的肺癌标本中,Rb2/p130基因突变的检出率为16.67%,其中肺腺癌的突变率为20%（4/20）,肺鳞癌的突变率16.67%（1/6）;在检测过程中发现PCR产物浓度和杂交温度是影响结果的关键因素,对突变位点的测序证实了基因芯片的结果。结论：基因芯片法检测出中国人肺癌组织标本存在Rb2/p130基因突变,但突变率（16.67%）低于西方人（78.5%）。未能检测全部突变位点、标本量小和人种差异可能是低突变率的原因。Objective：Part of the hot-spot mutations of Rb2/p130 gene was screened in patients with lung cancers by oligonucleotide gene chips in order to explore the rapid and accurate method in gene diagnosis of lung cancer. Methods： Surgical specimens of patients with lung cancer were collected and genomic DNAs were extracted. After DNA sequence of Rb2/p130 was analysed, probe and primer were synthesized,and gene chips were made by doting the probes to modified slide. PCR amplification of Rb2/p130 were performed using fluorescently labelled primers.Hybridization of the products of PCR amplification and gene chips which was used to detected whether having hot-spot mutation,and hybridization signal were detected in order to assess hot-spot mutation. Results：With oligonucleotide gene chips, we screened the Rb2/p130 gene mutation rate was 16.67% in 30 sample of lung cancer in which the mutation rate of lung adenocarcinoma was 20%（4/20）, the mutation rate of lung squamous cell carcinoma was 16.67%（1/6）.We conclude that the concentration of PCR production and hybridization temperature were the key factors affecting the results. Conclusion：Rb2/p130 mutations in surgical specimens can be detected by gene chips, but the percentage of mutation in Chinese lung cancer patients（16.67%）was lower than that of Occident lung cancer patients（78.5%）. The reasons of low mutation rate in Chinese lung cancer may contribute to be unable to detect all the mutation loci,too small sample size and the different human races.

关 键 词：肺癌 基因芯片 Rb2/p130基因 点突变 

分 类 号：R734.2[医药卫生—肿瘤] Q7[医药卫生—临床医学]