【作者】 郗雪艳； 杨秋霞； 李殿俊； 王辉； 张明宇； 王丽荣；

【Author】 XI Xue-yan, YANG Qiu-xia,LI Dian-jun, et al (Department of Immunology, Harbin Medical University, Harbin 150086, China)

【机构】 哈尔滨医科大学免疫学教研室；

【摘要】 目的获取肺腺癌及同源正常组织完整的RNA分子,并由此逆转录成全长cDNA,经各种杂交手段,筛选未知基因片段。方法从肺腺癌及同源正常组织中分别提取总RNA,经琼脂糖凝胶电泳确认RNA未降解后,在oli- godT-primer的引导下。将mRNA逆转录成ds cDNA,并在ds cDNA上游及下游连接上不同的适配子(adaptor),连接适配子的ds cDNA分别在不同的引物引导下,经PCR扩增并富集出大量的全长cDNA。结果扩增出肺腺癌及同源正常组织的全长cDNA,肺正常组织的cDNA基因片段在0．3-1．9kb之间,而肺腺癌的片段在0．3-4．5kb之间。结论长距离cDNA PER是从微量d8 cDNA起始材料中获得大量的全长cDNA信息的有效方法。更多还原

【Abstract】 Objective To obtain lung adenocarcinoma and homologous nonnal tissue intact RNA and tran scribe mRNA into full-length cDNA, from which screen unknown specific gene by hybridization. Methods We extracted total RNA from lung adenocarcinoma tissue and homologous normal tissue, after it was identified no degration by agar gel electrophorosis, transcribed mRNA into ds cDNA which was primed by oligodT-primer. Then ligated with two adaptor in upstream and downstream of ds cDNA respectively. Ds cDNA ligated with adaptor was amplified through Polymerase chain reaction. Results We amplified full-length cDNA of lung adenocarcinoma and homologous normal tissue. The cDNA fragments of normal tissue concentrated in 0.3 ~ 1. 9kb and cDNA fragments of tumor tissue concentrated in 0.3 - 4. 5kb. Conclusion Long distance cDNA PCR is an effective method to get magnanimous full-length cDNA information from a small quantity ds cDNA materials.更多还原