【作者】 郝权； 徐勇； 李文录； 赵小鸽； 刘文欣；

【Author】 HAO Quan XU Yong-Li Wen-Lu et al Pelvis department Tianjin Cancer Hospital affiliated to Tianjin Medical University Tianjin 300060 China

【机构】 天津医科大学附属肿瘤医院盆腔科； 天津医科大学第二医院泌尿科； 西安交通大学医学院医学实验中心；

【摘要】 目的:当对有间质细胞混杂的膀胱移行上皮细胞与癌细胞进行基因差异表达分析时,激光捕获显微切割技术 (LCM)是不可缺少的。LCM与RNA线性扩增结合,目的是确定一条可行的技术路线,获取均质的、足量的RNA,以备进一步用于膀胱癌相关基因研究。方法:采用LCM技术分别从正常膀胱粘膜及膀胱癌组织冰冻切片中获取膀胱移行上皮细胞及癌细胞,提取RNA,并对120ng上皮细胞RNA进行线性扩增,获得aRNA 20ug。用RT—PER验证扩增前、后 RNA中β-actin基因表达水平。结果:对照实验I证实 LCM后RNA完整性较好;产物RNA扩增后获得片段大小为 0．5—2．5kb的aRNA,且β-actin表达完整。结论:LCM 结合RNA线性扩增技术获取均一的、足量的、完整性好的目的细胞RNA,能用于进一步研究。更多还原

【Abstract】 Objective: Laser Capture Microdisection (LCM ) has become indispensable for the analysis of the difference of gene expression between human bladder transitional cell and bladder transitional cell carcinoma (BTCC). The study was designed to determine a feasible technical routine combining LCM with RNA Linear Amplification In vitro to obtain pure, sufficient, integrative RNA for the using in further research. Methods: Bladder transitional cell were obtained from frozen bladder membrane sections applying LCM, by the same taken, BTCC cells from frozen BTCC tissue section. Then RNA was extracted and about 120ng RNA was linearly amplified in vitro,then 20ug aRNA was obtained. The expression levels of β - actin in primary total RNA and amplified RNA were detected using RT-PCR. Results: RNA integrity is good after LCM confirmed by contral experiment I, About 0.5-2.5kb RNA fragments were obtained after RNA amplification and p - actin levels were integral. Conclusion: LCM combined with RNA Linear Amplification In vitro can be apllied to obtain pure, sufficient, integrative RNA for the using in further research.更多还原