利用基因芯片技术检测BRAF基因在胰腺癌组织中的表达  

Detection of expression of BRAF gene in pancreatic adenocarcinoma by oligonucleotide microarray

作　　者：汤永辉[1] 石欣[1] 卫文俊[1] 程张军[1] 高乃荣[1] 

机构地区：[1]东南大学附属中大医院普外科,江苏南京210009

出　　处：《东南大学学报（医学版）》2006年第5期345-348,共4页Journal of Southeast University(Medical Science Edition)

基　　金：国家自然科学基金(30500491);教育部留学归国人员科研启动基金(6890001004);江苏省卫生厅科研项目基金(7690008236)资助项目

摘　　要：目的：研究BRAF基因在正常胰腺与胰腺癌组织中表达的情况。方法：收集正常胰腺组织3例以及胰腺癌手术切除标本6例。后者均经病理学检查证实；用TRizol法对组织进行总RNA抽提，采用无RNA酶的DNA酶I等方法进行纯化；通过紫外分光光度计、琼脂糖凝胶电泳以及Lab-on—chip对总RNA进行质控；利用寡核苷酸芯片技术检测正常胰腺和胰腺癌组织中BRAF基因的表达，并且应用逆转录聚合酶链反应（RT‘PCR）技术加以验证。结果：总RNA的OD260nm／OD280nm之值在1．8～2．0之间；琼脂糖凝胶电泳18s和28s电泳条带清晰，且28s和18s亮度之比≥2；Lab—on—chip检测显示18s和28s双峰比例及位置均正常。无降解杂峰出现。芯片杂交扫描图像信号清晰，具有较低的整体背景和较高的信噪比；Ratio分析表明，BRAF基因在胰腺癌组织中表达上调（Cy3／Cy5〉2．0）。RT—PCR验证了胰腺癌组织中BRAF mRNA表达上调。结论：BRAF基因在胰腺癌组织中表达上调，其在胰腺癌发生发展过程中的作用机制还有待于进一步探讨。Objective To investigate the expression of v-raf murine sarcoma viral oncogene homolog B1 （BRAF） gene in normal pancreas and pancreatic adenocarcinoma tissue. Methods Three cases of normal pancreas tissue and six cases of pancreatic adenocarcinoma were collected, which were confirmed by pathology. The RNA was extracted through the method of TRizol, purified by DNA enzyme I and detected through violet-spectrophotometer, agarose gel electrophoresis （AGE） and Lab-on-chip methods. The expression of BRAF gene was detected by oligonucleotide microarray,which was comfirmed by reverse transcription polymerase chain reaction （RT-PCR）. Results The ratio of OD260nm/OD280nm was between 1.8 and 2. O. The bands in 18 s and 28 s through AGE was clear, and the brightness ratio of the two bands （28 s and 18 s） was more than 2. O. The scales and situs of split-blip were normal through the detection of Lab-on-chip method. BRAF mRNA was overexpressed in pancreatic cancer tissue compared with normal pancreas. The scanning signal of microarray hybridization was clear, and the images had a lower background and higher signalnoise ratio. The expression of BRAF gene in pancreatic adenocarcinoma against normal pancreas was up-regulated（ Cy3/ Cy5 〉 2. 0）by Ratio analysis. Conclusion BRAF gene is overexpressed in pancreatic adenocarcinoma and may be involved in the pathogenesis of pancreatic cancer, but the role BRAF gene in the mechanisms of pancreatic cancer should be further investigated.

关 键 词：胰腺癌 BRAF基因 寡核苷酸芯片 逆转录聚合酶链反应 

分 类 号：R735.9[医药卫生—肿瘤] R-33[医药卫生—临床医学]