Key Laboratory of Genetics and Biotechnology, College of Life Science, Capital Normal University, 100048 Beijing, China.
J Agric Food Chem. 2010 Mar 10;58(5):2777-86. doi: 10.1021/jf903363z.
High molecular weight glutenin subunits (HMW-GS) from 60 germplasms including 30 common wheat cultivars and 30 related species were separated and characterized by a suite of separation methods including sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), reversed-phase high-performance liquid chromatography (RP-HPLC), high-performance capillary electrophoresis (HPCE), and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS). Comparative analysis demonstrated that each methodology has its own advantages and disadvantages. The main drawback of SDS-PAGE was its overestimation of molecular mass and incorrect identification of HMW-GS due to its low resolution. However, it had the advantages of technical simplicity and low requirements of equipment; thus, it is suitable for large-scale and high-throughput HMW-GS screening for breeding programs, especially when the glutenin composition is clear in the breeding material. MALDI-TOF-MS clearly expressed many technical advantages among the four methods evaluated, including high throughput, high resolution, and accuracy; it was, however, associated with high equipment cost, thus preventing many breeding companies from accessing the technology. RP-HPLC and HPCE were found to be intermediate between SDS-PAGE and MALDI-TOF-MS. Both RP-HPLC and HPCE demonstrated higher resolution and reproducibility over SDS-PAGE but lower detection power than MALDI-TOF-MS. Results demonstrated that MALDI-TOF-MS is suitable for analyzing HMW-GS for routine breeding line screening and for identifying new genotypes.
利用十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）、反相高效液相色谱（RP-HPLC）、高效毛细管电泳（HPCE）和基质辅助激光解吸/电离飞行时间质谱（MALDI-TOF-MS）等一系列分离方法，对 60 份种质资源（包括 30 个普通小麦品种和 30 个相关物种）的高分子量谷蛋白亚基（HMW-GS）进行了分离和鉴定。比较分析表明，每种方法都有其自身的优点和缺点。SDS-PAGE 的主要缺点是分辨率低，导致其对高分子质量的估计过高，对 HMW-GS 的鉴定不正确。然而，它具有技术简单、设备要求低的优点；因此，它适用于大规模高通量的 HMW-GS 筛选，特别是在育种材料中谷蛋白组成清晰的情况下。MALDI-TOF-MS 在评估的四种方法中明显表现出许多技术优势，包括高通量、高分辨率和准确性；然而，它与设备成本高有关，因此许多育种公司无法获得这项技术。RP-HPLC 和 HPCE 被发现介于 SDS-PAGE 和 MALDI-TOF-MS 之间。RP-HPLC 和 HPCE 均表现出比 SDS-PAGE 更高的分辨率和重现性，但检测能力低于 MALDI-TOF-MS。结果表明，MALDI-TOF-MS 适用于常规育种系筛选和新基因型鉴定中的 HMW-GS 分析。
