Identification Of Seed Storage Proteins And Molecular Cloning Of Gliadins In Wheat

As major components of wheat storage proteins, the composition and content of wheat glutenins and gliadins play an important role in wheat quality. Separation and characterization of seed storage proteins and directed clone of their encoding genes are highly important for the study of gluten structure and the related relationship with quality, which can also provide alternative resources for improvement of wheat quality through gene engineering.The study of wheat storage proteins grows with the development of separation technology. As the invention of advanced separation techniques, it is possible to study the structures of wheat seed proteins on molecular level. For instance, reversed-phased high performance liquid chromatography (RP-HPLC) and mass spectrophy (MS) provides us fast and accurate tools to analyze wheat protein samples on a large scale.· Separation and identification of wheat storage proteins by RP-HPLCIn this study, the factors influencing RP-HPLC separation, such as column temperature, elution grads, sample volume, extract time of sample and inject volume of sample, were optimized to establish RP-HPLC analysis system for separating wheat gliadins and glutenins. The technical system established included that Agilent 1100 instrument and reversed-phased colomn (ZORBAX 300SB-C18 Stable Band Analytical 4.6 × 250mm, 5-Micron) were used. The major analytical parameters were: column temperature 50℃, flow speed 1.00ml/min, sample volume 5μl, eluting grade and the variable concentrations of acetonitrile growing from 21% to 48% (v/v) in 55 minutes. The result shows that the HMW-GS 1 Ax1, 1Bx6, 1Bx7, 1Bx13, 1Bx14, 1Bx17, 1Dx2, 1Dx5, lBy8, lByl8, 1Dy10 and 1Dy12 could be well separated while the HMW-GS 1Dx2 and 1By15, 1Dx5 and 1By15, 1Dx2 and 1By20, 1Dx5 and 1By9, 1Dx5 and 1By18, 1Dx2 and lByl6 could not be separated by this method. In addition, a procedure for the identification of wheat gliadin γ45 relating to good quality was established using the above technical system, which could provide an alternative choice to identify wheat quality.· Identification of wheat gliadins by MALDL-TOF-MSThe accurate measurement of molecular weights and characterization of wheat gliadins through matrix assisted laser desorption ionization time of flight mass spectrometry...