Establishment and Application of Multiplex PCR Systems Based on Molecular Markers for HMW-GSs in Wheat

Abstract

High-molecular-weight glutenin subunits (HMW-GSs) encoded by alleles at the Glu-A1, Glu-B1, and Glu-D1 loci confer unique end-use quality properties of common wheat (Triticum aestivum L.). Wheat accessions with the high-quality HMW-GSs combination of Ax2*/Bx7OE/Dx5 usually exhibit strong gluten characteristics. In order to stack these three high-quality subunit genes by molecular markers in strong gluten wheat breeding, an agarose gel-based multiplex PCR marker for these high-quality HMW-GSs and two agarose gel-based multiplex PCR markers detecting the homozygosity of Ax2* and Bx7OE subunits were developed. These markers were verified in an F2 segregating population from a cross between a medium-gluten winter wheat cultivar with the HMW-GSs combination of Ax null/Bx7 + By8/Dx4 + Dy12 and a strong-gluten spring wheat cultivar with the HMW-GSs combination of Ax2*/Bx7OE + By8*/Dx5 + Dy10. By integrating the newly established multiplex PCR markers and a published co-dominant PCR marker of the Dx5 subunit, a complete molecular marker selection system was established. After multiple rounds of molecular marker-assisted selection with the system, 17 homozygous winter wheat lines that stacked the three high-quality HMW-GSs were generated. The gluten strength of these homozygous lines was comparable to their strong-gluten parent, but significantly higher than that of their medium-gluten parent by measuring their lactic acid-sodium dodecyl sulfate solvent retention capacities of whole wheat meal. The multiplex PCR systems established in the present study can be used for molecular marker-assisted selection of strong gluten wheats.