Establishment of two-dimensional gel electrophoresis for soybean protein isolate and its application
2020
Lu, Xinkai | Cui, Yaoming | Guan, Junjun | Liu, Xue | Zhu, Hao | Ji, Xuyang | Zheng, Jianzhang | Cheng, Yunlong | Fu, Xiaofei
To optimize the conditions for the establishment of two-dimensional gel electrophoresis (2-DE) of soybean protein isolate (SPI), we investigated Ampholine mixture, anodic and cathodic electrolytes, loading amount of sample, acrylamide concentration, pH gradient and gel staining method in two-dimensional gel electrophoresis to optimize the protein imaging conditions in two-dimensional gel. The results of mixed-level design experiments showed that Ampholine, loading amount and gel staining method had significant effect (P < 0.05) on 2-DE of SPI. The optimal conditions were Ampholine mixture (pH 3–10 + pH 5–7 or pH 4–6 + pH 5–7), loading amount of 2 mg sample and silver staining. Although the acrylamide concentration of the gel, the pH gradient, the anodic and cathodic electrolyte solutions had significant statistical effects on the protein separation degree, the complexity of the protein composition and the visibility of the gel images were more inclined to the 12% gel, the 3–10 pH gradient and the H₃PO₄/NaOH electrolyte. According to the established conditions, the hydrolyzed products of SPI emulsion were determined by 2-DE, and the dynamic changes of protein in the process of enzymatic hydrolysis were described.
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