Low molecular weight proteins are promising molecules for rice plants. The objectives of this study were to develop an optimized method for protein extractions and protein chips and to perform practical application for isolation of low molecular weight proteins/peptides as potential biomarkers of three rice cultivars, Ilpumbyeo, Hwaseongbyeo, and Nakdongbyeo, using surface-enhanced laser desorption/ionization (SELDI) time-of-flight (TOF) mass spectrometry (MS). Among five protein extraction methods tested, we found that the trichloroacetic acid (TCA)-acetone method was the most effective for the extraction of low molecular weight proteins/peptides. Four protein chip arrays, including CM10 (weak cation exchanger), Q10 (strong anion exchanger), H50 (reverse phage), and IMAC30 (immobilized metal affinity), were used under low and high stringency conditions at varying pH with two energy absorbing molecules, alpha-Cyano-4-hydroxycinnamic acid (CHCA) and sinapinic acid (SA). Of the 70 proteins isolated by the Q10 array, 20 showed significant differences in expression between the three cultivars (p < 0.05). Principal component analyses showed distinct expression patterns in the three cultivars, which were confirmed by hierarchical cluster analysis-based heat mapping. Significantly upregulated proteins were divided into five cultivar-specific clusters by heat mapping. Taken together, the results indicate that rice cultivar-specific small peptides that may be the potential biomarkers have been proposed by the TCA-acetone extraction method in combination with the Q10-SA chip array.

低分子量蛋白质是水稻植物的有前途的分子。本研究的目的是开发蛋白质提取和蛋白质芯片的优化方法，并在实际应用中分离低分子量蛋白质/肽作为三个水稻品种 Ilpumbyeo、Hwaseongbyeo 和 Nakdongbyeo 的潜在生物标志物，使用表面增强激光解吸/电离 (SELDI) 飞行时间 (TOF) 质谱 (MS)。在测试的五种蛋白质提取方法中，我们发现三氯乙酸 (TCA)-丙酮方法对于提取低分子量蛋白质/肽最有效。四种蛋白质芯片阵列，包括CM10（弱阳离子交换剂）、Q10（强阴离子交换剂）、H50（反向噬菌体）和IMAC30（固定化金属亲和力），在不同 pH 值的低和高严格条件下使用两种能量吸收分子，α-氰基-4-羟基肉桂酸 (CHCA) 和芥子酸 (SA)。在 Q10 阵列分离的 70 种蛋白质中，有 20 种在三个栽培品种之间显示出显着的表达差异（p  < 0.05)。主成分分析显示三个品种中不同的表达模式，这通过基于层次聚类分析的热图得到证实。通过热图将显着上调的蛋白质分为五个品种特异性簇。综上所述，结果表明，通过 TCA-丙酮提取方法结合 Q10-SA 芯片阵列，已经提出了可能是潜在生物标志物的水稻品种特异性小肽。