Characterization of High Mr Subunits of Glutenin by Combined Chromatographic (RP-HPLC) and Electrophoretic Separations and Restriction Fragment Length Polymorphism (RFLP) Analyses of their Encoding Genes

Abstract

A comparison was made of the electrophoretic (SDS-PAGE) and reversed-phase high-performance liquid chromatographic (RP-HPLC) separations of several allelic variants of high Mr glutenin subunits from durum and bread wheats. The results showed that the order of elution of subunits Dy, By, Dx, Bx and Ax on RP-HPLC was inversely related to their mobilities in SDS-PAGE. Subunits having similar apparent Mrs and isoelectric points had different surface hydrophobicities. The chromatographic results highlighted similarities between the structural genes encoding x- or y-type subunits present in different chromosomes (homoeallelic genes) and differences between x and y subunits present on the same chromosome, confirming previous results obtained by SDS-PAGE analyses. Differences in elution times that were observed when the same subunit was either reduced or reduced and alkylated with 4-vinylpyridine were related to the number cysteine residues and to the specific genome containing the corresponding encoding gene. Restriction fragment length polymorphism (RFLP) studies indicated that some subunits, which had higher apparent Mrs, produced larger fragments; the absence of some subunits was shown to be due to deletion or lack of expression of their corresponding encoding genes.