Wheat cultivar-specific proteins in grain revealed by 2-DE and their application to cultivar identification of flour

Wheat flour proteins were studied to identify the cultivar‐specific proteins and use them to identify cultivars in flours. Proteins extracted from flours of Japanese wheat (cultivars Hokushin, Horoshirikomugi, Kitanokaori and Kachikei 33) and Canadian wheat (Canada Western Red Spring Wheat No. 1; 1CW) were analyzed by 2‐DE with IEF gels over three pH ranges: pH 4–7, pH 5–8, and pH 6–11. This system enabled detection of more than 1600 protein spots. We recognized that among 50 protein spots showing cultivar‐dependent qualitative changes, 25 proteins were wheat cultivar specific. These 50 protein spots were analyzed by N‐terminal Edman degradation microsequencing and MALDI‐TOF‐MS; 21 protein spots were storage proteins, such as gliadin and low‐molecular mass glutenin subunit. Five protein spots were identified as dehydroascorbate reductase (Triticum aestivum), triticin precursor (T. aestivum), alpha‐amylase inhibitor (Oryza sativa), DNA‐binding with one finger (Dof) zinc family protein (O. sativa), and nonphototropic hypocotyl 1 (NPH1) protein (Avena sativa). The other protein spots appeared to be hypothetical proteins (O. sativa or Arabidopsis thaliana) or functional unknown proteins. These specific proteins can be used as markers to identify wheat cultivars in blended flour composed of two or three flours.