Two y-type high molecular weight glutenin subunits (HMW-GSs) 1Ay12* and 1Ay8* from the two accessions PI560720 and PI345186 of cultivated einkorn wheat (Triticum monococcum ssp. monococcum, AA, 2n=2x=14), were identified by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The mobility of 1Ay12* and 1Ay8* was similar to that of 1Dy12 and 1By8 from common wheat Chinese Spring, respectively. Their ORFs respectively consisted of 1812 bp and 1935 bp, encoding 602 and 643 amino acid residues with the four typical structural domains of HMW-GS including signal peptide, conserved N-, and C-terminal and central repetitive domains. Compared with the most similar active 1Ay alleles previous published, there were a total of 15 SNPs and 2 InDels in them. Their encoding functions were confirmed by successful heterogeneous expression. The two novel 1Ay alleles were named as 1Ay12* and 1Ay8* with the accession No. JQ318694 and JQ318695 in GenBank, respectively. The two alleles were classed into the two distinct groups, Phe-type and Cys-type, which might be relevant to the differentiation of Glu-A1-2 alleles. Of which, 1Ay8* belonged to Cys-type group, and its protein possessed an additional conserved cysteine residue in central repetitive region besides the six common ones in N- and C-terminal regions of Phe-type group, and was the second longest in all the known active 1Ay alleles. These results suggested that the subunit 1Ay8* of cultivated einkorn wheat accession PI345186 might have a potential ability to strengthen the gluten polymer interactions and be a valuable genetic resource for wheat quality improvement.
Fig. 1. 1Ay subunits, and their encoding gene DNA amplification and heterologous expression of T. monococcum ssp. monococcum.
A: SDS-PAGE profile of the two accessions PI560720 (lane 2) and PI345186 (lane 3) and the reference Chinese Spring (line 1), 1Ay12* and 1Ay8* indicate their 1Ay subunits, respectively;
B: the amplified complete ORF of 1Ay12* (lane 1) and 1Ay8* (lane 2) by PCR;
C and D: bacterial expression of the modified ORFs of 1Ay12* and 1Ay8* in E. coli, respectively, with lane 1 being the HMW-GSs from PI560720 (C) and PI345186 (D), lanes 2 (the selective extract) and 3 (the general extract) being the expressed 1Ay proteins in IPTG-induced bacterial cells, lane 4 being the total proteins in the bacteria cells without IPTG induction for control,
and the expressed proteins of 1Ay12* and 1Ay8* were respectively marked by arrow.
1. Author Information:
Xiao-Hui Guo, Bi-Hua Wu, Xi-Gui Hu, Zhe-Guang Bi, Zhen-Zhen Wang,Deng-Cai Liu, You-Liang Zheng
Correspondence: E-mail: wubihua2001@yahoo.com.cn (B.-H. Wu).
2. Publication History: Gene 516 (2013) 1–7