Molecular characterization of novel high-molecular-weight glutenin subunit genes from Aegilops tauschii

The <i>Glu-D1</i> locus is known to contribute the most to gluten quality in common wheat. <i>Aegilops tauschii</i>, as a donor species of the common wheat D genome, is of great significance for improving wheat flour processing quality by exploring and utilizing its excellent quality-related genes. In this study, high-molecular-weight glutenin subunit (HMW-GS) compositions of 173 <i>Aegilops tauschii</i> accessions from the Middle East, Central Asia, and Xinjiang were analyzed using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) technology. A total of 13 subunit types were detected at the G<i>lu-D1 ^t</i> locus, including the unique subunit types absent in common wheat. Unexpectedly, all these <i>Ae. tauschii</i> accessions lacked the extra cysteine residue at position 118 in their Glu-1Dx subunits detected by a specific functional marker, <i>dCAPS5</i>. To determine whether other sites of Glu-D1^t subunits in <i>Ae. tauschii</i> contains the extra cysteine residue, the grain RNA of <i>Ae. tauschii</i> accessions with rare subunit types at different days post-anthesis (6th, 9th, 12th, 15th, 18th, 21st, and 24th days) were mixed and sequenced using third-generation (PacBio) transcriptome sequencing technology. A total of 12 new HMW-GS genes were obtained, including 6 genes each for <i>Glu-D1x</i> and <i>Glu-D1y</i>. Sequence alignment analysis revealed the extra cysteine residue was not observed in the protein sequences deduced by 12 new HMW-GS genes. Interestingly, HMW-GS of <i>Ae. tauschii</i> accessions AY16 and AY49 had longer repetitive domain insertions, while those of AY16 and AY53 exhibited higher <i>α</i>-helix and β-strand contents, which may positively influence dough properties. Additionally, the relative content of HMW-GS in AY11, AY32, and AY49 was significantly higher than that of the <i>Glu-D1</i> subunits in wheat variety Xinmai 26 with the strong gluten quality, favoring the formation of larger glutenin polymers. From this point of view, <i>Ae. tauschii</i>, as a potentially valuable germplasm resource, can provide rich subunit gene resources for quality breeding in common wheat.