小麦醇溶蛋白毛细管电泳分离技术及其在品种鉴定上的利用研究

Studies on Capillary Electrophoresis Separation of Wheat Gliadins and Its Use in Varietal Identification

本文探讨了毛细管电泳(CE)技术分离小麦醇溶蛋白的最佳条件及用于品种鉴定的潜力.大量检测结果发现,以下分离条件可获得较高的分辨度和重复性,即CE缓冲液用碱性硼酸钠(pH9.0,并含20%乙腈和1%SDS),毛细管长度47cm,内径50μm,运行电压15kV,毛细管温度30℃.在这些条件下,一个品种在25～30分钟之内,可分离出35个以上的醇溶蛋白组分(峰或峰肩),而且重复性较高,峰迁移时间的偏差最高值在0.03分之内.同一品种在不同年度和不同生态条件下,也表现一致的醇溶蛋白CE模式.17个小麦品种的CE图谱均显示出明显的质量和数量上的差异.其中一些亲缘关系极近的品种,其醇溶蛋白聚丙烯酰胺凝胶电泳(PAGE)图谱相同,但它们的CE图谱则存在显著差异.因此,毛细管电泳是一项分辨度高、分离速度快、重复性好、自动化程度高的蛋白质分离新技术,在小麦品种鉴定中具有十分诱人的应用前景.

The optimal conditions of gliadin capillary electrophoresis(CE) and potential use for varietal identification were studied in this article. Gliadin proteins were extracted with 30% aqueous ethanol. CE was performed on a new Beckman P/ACE 5500 system with Gold Software for system control and data acquisition. High resolution and reproducibility of gliadin electrophore-grams were achieved by using 47 cm length(50 μm i. d. ) capillary, 15 kV at 30℃ in 0. 06 M sodium borate buffer(pH9. 0), containing 20% acetonitrile and 1% SDS. Using the conditions, one cultivar can be separated into more than 35 gliadin components (peaks or shoulders) in 25～30 min. Gliadin CE patterns of 17 winter wheat cultivars showed apparent qualitative and quantitative differences. In particular, some closely related cultivars that their gliadin A-PAGE elec-trophoregrams are identical, can also be easily discriminated by using CE. As shown in thisstudy, capillary electrophoresis appears to be capable of rapid, high-resolution and reproducible, automated separation of gliadins, and has great potential for wheat varietal identification.