| High molecular weight (HMW) glutcnin subunits, as one of the seed storage proteins in wheat, its composition and amount have a profound influence on the baking quality of wheat. The number of functionally superior HMW glutenin subunits identified in common wheat is veiy limited. The Aegilops genus is veiy closely related to the common wheat The types of HMW glutenin subunits in Aegilops species are much richer than those in bread wheat For the better understanding of the molecular structure difference between HMW glutenm subunits specified by the Aegilops species and those of wheat and their potential utilization for the wheat quality improvement, the HMW glutenin subunits composition of 51 accessions from Aegiiops tawchii(DD,2n=2x= 14) and and I accession from Aegilops caudata were studied using SDS-PAGE and Western-blotting. In addition, six novel IIMW glutcnin genes were isolated and expressed in E. coil.1.11MW ghiteuin subunit compsition analysis of AqUops tausckia?Two novel types of 11MW glutenin subunits were observed from 51 accessionsof Aegilops tawchii , which were designated as Dx2. It and Dy 13t respectively. Thefrequencies of 7 subunit combinations(2+12,5+10,2+10,5+12, 2.lt +10, 2.1?+12 and2+ 13t) are 27.5/o(2+ 12), 25.5扽o(5+ 10), 31 .4/o(2+ 1O),3.9/o(5+ 12), 2.0/o(2. It +10),2.0% (2.1?+12) and 7.8%( 2+13t) respectively.2. Molecular cloning of Dyl3 subunit gene from Aegilops t.ausckliThe complete nucleotide sequence of the Dy13?subunit contained 1878 base parirs. The deduced amino acid sequence of Dy13?contained a singnal peptide of2l amino acids, a N-terminal region of 104 amino acids, a C-terminal region of 4293amino acids and a central repetitive domain of 457 amino acids. There were 47 hcxapcptides (PGQGQQ), 17 nonapeptides (GYYPTSLQQ), two peptides containin& respectively, 12(GYYPSVTSPRQG), 1 1(SYYPGQASPQQ) and a tripeptide (GYD) in the repetitive domain. Among all the known 11MW glutenin subunits encodeed by D-genome, the coding region of Dy13?was the shortest Its sequences is obviously different from DyIO and Dy 12. Therefore , Dy 13t is a novel subunit characterized and has an itnportant utilization in the wheat quality improvement.3. Molecular doming of Dx2.1?subunit from AqIIeps taaasckliThe complete nucleotide sequence of the Dx2. it gene contained 2514 base pairs. The deduced amino acid sequence of Dx2.lt contained a singnal peptide of2i amino acids, a N-terminal region of 89 amino acids, a C-tcrminai region of42 amino acids and a central repetitive domain of 684 amino acids. Exception for hexapeptidcs (PGQGQQ) and nonapcptides (GYYPTSLQQ), many additional tripeptides(GQQ) were detected in the repetitive domain.4. Molecular doming of Cx and Cy subunit from Aqilops caudataThe complete nuclcotide sequence of the Cx and Cy genes contained 2391 and 1905 base paires. The proteins encoded by these two genes consisted of signal peptide, a non-repetitive N-tcnninal and a C-terminal region, flanked by a repetitive domain. The great differences in amino acid composition and order of hexapeptides and nonapeptides between Cx and Cy subunits and other 11MW subunits in bread wheat indicated that they are new members of HMW glutenin family.5. Phylogeuetic relationships of Dyl3t, j)~1t Cx and CyIn the Phylogenetic tree map,Dyl3t is veiy closely related to the y-type genes encoded by D genome of common wheat. Although DylO,Dy12,Dyl2t,and Dyl3t are all encoded by the D genome ,the Dyl3tis more closed related to the DyI2t than both DylO and Dy12. Dx2.1?is more closely clustered to both Dx2 and Dx5. The Cx is closely associated with D genome subunits compared with R and A94genome, while Cy is very closely related to the Cy from Aegilops cylindrica(2n=4xz=28,CCDD).6. Expression of High molecular weight glutenin subunits in E. coilThree high molecular weight gluteninsubunits (Dx2.lt, Cx, Cy) have been successfully expressed in E.coli (BL21(DE3)plsS. The expressed HMW glutenin subunits of cloned genes in E... |
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