Identification of new low-molecular-weight glutenin subunit genes in wheat

Abstract

To clarify the composition of low-molecular-weight glutenin subunits (LMW-GSs) in a soft wheat cultivar, we cloned and characterized LMW-GS genes from a cDNA library and genomic DNA in Norin 61. Based on alignment of the conserved N- and C- terminal domains of the deduced amino-acid sequences, these genes are classified into 12 groups. One of these groups (group 5), the corresponding gene of which has not been reported previously, contains two additional hydrophobic amino-acid clusters interrupting the N-terminal repetitive domain. Other groups (groups 11 and 12), which were not identified in other cultivars as a protein product, showed all eight cysteines in the C-terminal conserved domain. With specific primer sets for these groups it was revealed that Glu-D3 and Glu-A3 encoded the former and the latter, respectively. Both groups of genes were expressed in immature seeds. The presence of these groups of LMW-GSs may affect the dough strength of soft wheat.

Molecular and Immunological Characterization of Tri a 36, a Low Molecular Weight Glutenin, as a Novel Major Wheat Food Allergen

“…Immature seeds were harvested 7,10,15,20,25,30, and 35 d after onset of pollination directly into liquid nitrogen and stored at 280˚C until use (20). Rye, barley, oat, spelt, and rice grains produced by Alnatura (Bickenbach, Germany) were bought at a local drugstore.…”

Characterization of wheat gluten proteins by hplc and MALDI TOF mass spectrometry

“…The DNA sequences for the group 3 type II LMW subunits show very strong similarities [20], so the peptide mass maps should be expected to have significant overlap. As a result, the data do not provide definitive evidence that all of the expected subunits are present.…”

“…The presence of BAB78741 and BAB78742 are perhaps the most definitive, since they both have one predicted peptide not present in the others. For© the© tentatively© identified© LMW© group© 2© type© I subunit© (BAB78738),© three© of© the© six© predicted© peptides were© observed© in© the© MALDI© QqTOF© peptide© pattern (data©not©shown).©A©higher©mass©peptide©was©also identified©at©5426.3©Da,©which©matches©a©predicted peptide© at© 5426.2© Da.© Now,© only© a© few© DNA© sequences for© this© group© have© been© determined© in© contrast© to© the group© 3© subunits© discussed© above© [20].…”

Identification of Specific Low-Molecular-Weight-Glutenin Subunits Related to Gluten Quality in Bread Wheats

“…PCR reactions were performed in a total volume of 25 µL containing 1.5 mM MgCl 2 , 0.1 mM of each dNTP, 5 pmol of Glu13 (5'-TTGGGGCTGT-TGTTGCTGATA-3') and Glu22 (5'-CGTCTTTGC-CCTCCTCGCTC-3') primer set 9 and 0.5 U AmpliTaq Gold DNA polymerase (Applied Biosystems Co., Ltd. USA), × 1 PCR Gold buffer and about 50 ng of total DNA. Reactions were performed according to the following protocol: denaturation at 94°C for 3 min; 35 cycles of 94°C for 30 s, 55°C for 30 s, 72°C for 90 s; and a final extension at 72°C for 5 min.…”

“…Cloned sequences of the LMW-GS genes revealed that all of the predicted LMW-GS amino acid sequences contained 8 cysteine residues 3,4 , which are considered to be involved in the formation of intra-and inter-molecular disulfide bonds. In our previous study, we analyzed the composition of the LMW-GS genes comprehensively in a soft wheat cultivar Norin 61 and classified these genes into 12 groups based on their deduced amino acid sequence similarity 9 . In this study, we analyzed the relationships between the LMW-GS composition and gluten strength (gluten index) using single-seed descent lines derived from the Kitakeiharu 717 (KKS 717) / Grandin cross.…”