Guttman Andras, Shi Liang, Khandurina Julia, Wang Xun
Torrey Mesa Research Institute, 3115 Merryfield Row, San Diego, CA 92121, USA.
J Chromatogr A. 2003 Oct 3;1014(1-2):29-35. doi: 10.1016/s0021-9673(03)00777-5.
Differential display analysis by cDNA fractionation, collection of differentially expressed fractions of interests and their downstream characterization is demonstrated. cDNA pools from two strains of Cochliobolus heterostrophus fungus were generated by specific restriction digestion and selective ligation. Micropreparative separation and isolation of differentially expressed transcript representatives were accomplished by high-performance capillary gel electrophoresis. The collected individual DNA molecules were polymerase chain reaction amplified and sequenced to create expressed sequence tags for the genes of interests. High resolving power and sensitivity of capillary gel electrophoresis enabled fast and automated processing of minute amounts of cDNA samples with high precision.
展示了通过cDNA分级分离进行差异显示分析、收集感兴趣的差异表达组分及其下游表征。通过特异性限制性消化和选择性连接产生了来自两种玉米小斑病菌株的cDNA文库。通过高效毛细管凝胶电泳完成了差异表达转录本代表的微量制备分离和分离。对收集到的单个DNA分子进行聚合酶链反应扩增和测序，以创建感兴趣基因的表达序列标签。毛细管凝胶电泳的高分辨率和灵敏度使得能够以高精度快速且自动化地处理微量cDNA样品。
