Characterization of high molecular weight glutenin subunit genes from the Ns genome of Psathyrostachys juncea

The Ns genome of the genus Psathyrostachys possesses superior traits useful for wheat improvement. However, very little is known about the high molecular weight (HMW) subunits of glutenin encoded by the Ns genome. In this paper, we report the isolation of four alleles of HMW glutenin subunit gene from Psathyrostachys juncea. Sequence alignment data shows the four alleles have similar primary structure with those in wheat and other wheat-related grasses, with some unique modifications. All four sequences more closely resemble y-type, rather than x-type, glutenins. However, our results show three of the subunits (1Ns2-4) contain an extra glutamine residue in the N-terminal region not found on typical y-type subunits, as well as the x-type subunit specific sequence LAAQLPAMCRL. These three subunits likely represent an intermediate state in the divergence between x- and y-type subunits. Results also indicate that the Ns genome is more closely related to the St genome of Pseudoroegneria than any other Triticeae genomes.     

长穗偃麦草中E和E1基因组高分子量麦谷蛋白基因启动子部分序列的进化分析

通过PCR克隆的方法,获得了分别来自二倍体长穗偃麦草的E基因组和四倍体长穗偃麦草的E_1基因组的4个高分子量麦谷蛋白亚基(HMW-GS)基因启动子的部分序列。序列分析表明,它们之间的同源性较高,两个x型亚基启动子序列之间只有1个碱基的差异,而两个y型亚基启动子序列完全相同,x和y型亚基启动子序列之间的长度和部分碱基位点都有差异。推测四倍体长穗偃麦草中的E_1基因组可能起源于二倍体的E基因组。与来自小麦族的A、B、D和G基因组部分亚基基因的启动子序列比较表明,小麦族的这一区域在进化上是相当保守的,不同基因组来源的序列同源性都在90％以上。经过对这些序列的聚类分析,表明长穗偃麦草的y型HMW-GS基因与其他亚基基因的进化关系较远,而x型亚基基因与一个来自小麦1B染色体的亚基基因关系最近。     

Characterization of HMW prolamines and their coding sequences from Crithopsis delileana

The high-molecular-weight (HMW) prolamines subunits and their coding sequences from wheat-related diploid species Crithopsis delileana were investigated. Only one HMW prolamine subunit with the similar electrophoresis mobility to the y-type HMW glutenin subunit of hexaploid wheat was observed in two accessions of C. delileana by SDS-PAGE analyses of the total storage protein fractions. It was confirmed by sequencing and expression analysis that this prolamine subunit was an x-type subunit. The amino acid sequence of this subunit had the similar typical structure to those of x-type HMW glutenin genes previously described in wheat. An in-frame stop codon was found in the coding sequences of y-type prolamine subunits. It was found by specifically extraction of HMW prolamines and sequence analysis that the coding regions of Ky prolamine subunit gene is very likely to be not expressed as a full-length protein. Phylogenetic analysis indicated that the Kx subunit could be clustered together with 1Ax1 subunit by an interior paralleled branch, and Ky subunit (inactive) was most closely related to the 1Ay subunit. The coding sequences of Kx subunit could successfully be expressed in bacterial expression system, and the expressed protein had the same electrophoresis mobility as the Kx subunit from the seed of C. delileana. It was the first time that the HMW prolamines subunits encoded by K genome of C. delileana were characterized.     

Characterization of two HMW glutenin subunit genes from Taenitherum Nevski

The compositions of high molecular weight (HMW) glutenin subunits from three species of Taenitherum Nevski (TaTa, 2n = 2x = 14), Ta. caput-medusae, Ta. crinitum and Ta. asperum, were investigated by SDS-PAGE analysis. The electrophoresis mobility of the x-type HMW glutenin subunits were slower or equal to that of wheat HMW glutenin subunit Dx2, and the electrophoresis mobility of the y-type subunits were faster than that of wheat HMW glutenin subunit Dy12. Two HMW glutenin genes, designated as Tax and Tay, were isolated from Ta. crinitum, and their complete nucleotide coding sequences were determined. Sequencing and multiple sequences alignment suggested that the HMW glutenin subunits derived from Ta. crinitum had the similar structures to the HMW glutenin subunits from wheat and related species with a signal peptide, and N- and C-conservative domains flanking by a repetitive domain consisted of the repeated short peptide motifs. However, the encoding sequences of Tax and Tay had some novel modification compared with the HMW glutenin genes reported so far: (1) A short peptide with the consensus sequences of KGGSFYP, which was observed in the N-terminal of all known HMW glutenin genes, was absent in Tax; (2) There is a specified short peptide tandem of tripeptide, hexapeptide and nonapeptide and three tandem of tripeptide in the repetitive domain of Tax; (3) The amino acid residues number is 105 (an extra Q presented) but not 104 in the N-terminal of Tay, which was similar to most of y-type HMW glutenin genes from Elytrigia elongata and Crithopsis delileana. Phylogenetic analysis indicated that Tax subunit was mostly related to Ax1, Cx, Ux and Dx5, and Tay was more related to Ay, Cy and Ry.     

Characterisation and analysis of new HMW-glutenin alleles encoded by the Glu-R1 locus of Secale cereale

This work reports the molecular characterisation of new alleles of the previously reported Glu-R1 locus. Wheat lines carrying the chromosome substitution 1R(1D), rye cultivars and related wild species were analysed. Five new x-type and four y-type Glu-R1 glutenin subunits were isolated and characterised. The coding region of the sequences shows the typical structure of the HMW glutenin genes previously described in wheat, with the N and C-terminal domains flanking the central repetitive region. Tri-, hexa- and nona-peptides found in the central repetitive region of wheat glutenin genes were also present in the rye genes. Duplications and deletions of these motifs are responsible for allelic variation at the Glu-R1 locus. Orthologous genes (from different genomes) were more closely related than paralogous genes (x- and y-type), supporting the hypothesis of gene duplication before Triticeae speciation. Differences in the number and position of cysteine residues identified alleles which in wheat are associated with good dough quality. SDS proteins encoded by some characterised alleles were presumptively identified.     

类大麦属高分子量谷蛋白基因Kx的序列测定及表达

利用SDS-PAGE检测了2份类大麦属(Crithopsis delileana)材料的高分子量谷蛋白亚基组成,并对其中1份材料的x型亚基进行了克隆和测序。结果表明,2份材料具有完全相同的蛋白电泳图谱。在小麦的高分子量区域仅检测到一条蛋白质带,与小麦y型亚基的迁移率接近,但克隆测序表明其为x型高分子量谷蛋白亚基,其编码基因命名为Kx。Kx基因编码区序列长度为2052bp．编码长度为661个氨基酸残基的蛋白质,其序列具有典型的x型高分子量谷蛋白亚基的特征。Kx基因能在原核表达系统内正确表达,其表达蛋白与来源于种子中的Kx亚基的迁移率完全一致。Kx亚基与小麦属A、B和D,山羊草属C和U以及黑麦属R染色体组编码的高分子量谷蛋白亚基氨基酸序列非常相似,但在N和C保守区的氨基酸组成以及重复区长度上与它们存在明显差异。聚类分析可将Kx与Ax1聚类为平行的分支。由此可见,来源于C．delileana的Kx基因为一新的x型高分子量谷蛋白亚基基因。     

Dynamics of the evolution of orthologous and paralogous portions of a complex locus region in two genomes of allopolyploid wheat

Two overlapping bacterial artificial chromosome (BAC) clones from the B genome of the tetraploid wheat Triticum turgidum were identified, each of which contains one of the two high-molecular-weight (HMW) glutenin genes, comprising the complex Glu-B1 locus. The complete sequence (285 506 bp of DNA) of this chromosomal region was determined. The two paralogous x-type ( Glu-1-1 ) and y-type ( Glu-1-2 ) HMW-glutenin genes of the complex Glu-B1 locus were found to be separated by ca. 168 000 bp instead of the 51 000 bp separation previously reported for the orthologous Glu-D1 locus of Aegilops tauschii, the D-genome donor of hexaploid wheat. This difference in intergene spacing is due almost entirely to be the insertion of clusters of nested retrotransposons. Otherwise, the orientation and order of the HMW glutenins and adjacent genes were identical in the two genomes. A comparison of these orthologous regions indicates modes and patterns of sequence divergence, with implications for the overall Triticeae genome structure and evolution. A duplicate globulin gene, found 5' of each HMW-glutenin gene, assists to tentatively define the original duplication event leading to the paralogous x- and y-type HMW-glutenin genes. The intergenic regions of the two loci are composed of different patterns and classes of retrotransposons, indicating that insertion times of these retroelements were after the divergence of the two wheat genomes. In addition, a putative receptor kinase gene near the y-type HMW-glutenin gene at the Glu-B1 locus is likely active as it matches recently reported ESTs from germinating barley endosperm. The presence of four genes represented only in the Triticeae endosperm ESTs suggests an endosperm-specific chromosome domain.     

Analysis of HMW glutenin subunits and their coding sequences in two diploid Aegilops species

Considerable progress has been made in understanding the structure, function and genetic regulation of high-molecular-weight (HMW) glutenin subunits in hexaploid wheat. In contrast, less is known about these types of proteins in wheat related species. In this paper, we report the analysis of HMW glutenin subunits and their coding sequences in two diploid Aegilops species, Aegilops umbellulata (UU) and Aegilops caudata (CC). SDS-PAGE analysis demonstrated that, for each of the four Ae. umbellulata accessions, there were two HMW glutenin subunits (designated here as 1Ux and 1Uy) with electrophoretic mobilities comparable to those of the x- and y-type subunits encoded by the Glu-D1 locus, respectively. In our previous study involving multiple accessions of Ae. caudata, two HMW glutenin subunits (designated as 1Cx and 1Cy) with electrophoretic mobilities similar to those of the subunits controlled by the Glu-D1 locus were also detected. These results indicate that the U genome of Ae. umbellulata and the C genome of Ae. caudata encode HMW glutenin subunits that may be structurally similar to those specified by the D genome. The complete open reading frames (ORFs) coding for x- and y-type HMW glutenin subunits in the two diploid species were cloned and sequenced. Analysis of deduced amino acid sequences revealed that the primary structures of the x- and y-type HMW glutenin subunits of the two Aegilops species were similar to those of previously published HMW glutenin subunits. Bacterial expression of modified ORFs, in which the coding sequence for the signal peptide was removed, gave rise to proteins with electrophoretic mobilities identical to those of HMW glutenin subunits extracted from seeds, indicating that upon seed maturation the signal peptide is removed from the HMW glutenin subunit in the two species. Phylogenetic analysis showed that 1Ux and 1Cx subunits were most closely related to the 1Dx type subunit encoded by the Glu-D1 locus. The 1Uy subunit possessed a higher level of homology to the 1Dy-type subunit compared with the 1Cy subunit. In conclusion, our study suggests that the Glu-U1 locus of Ae. umbellulata and the Glu-C1 locus of Ae. caudata specify the expression of HMW glutenin subunits in a manner similar to the Glu-D1 locus. Consequently, HMW glutenin subunits from the two diploid species may have potential value in improving the processing properties of hexaploid wheat varieties.     

Characterization of a novel variant HMW-glutenin gene from Elymus canadensis

High molecular weight (HMW) glutenin subunits (GS) play a key role in the determination of end-use quality of wheat and other cereal crops. In this study, we report the isolation and characterization of both promoter region and ORF of novel HMW-GS allele 1St1.3 from a perennial Triticeae species, Elymus canadensis. The amino acid (AA) sequences of E. canadensis 1St1.3 were deduced as 434 aa. Its protein primary structure comprises a signal peptide with a conserved N-terminal domain, a central repetitive domain and a C-terminal domain. E. canadensis 1St 1.3 possesses several distinct characteristics which are different from those of wheat HMW-GSs. The N-terminal domains of E. canadensis 1St 1.3 resemble that of y-type subunits, while their C-terminal domains are more similar to x-type subunits. The deletion of 85 bp fragment has been observed in promoter region of 1St 1.3, however which has not interrupted the expression of this gene. Our results indicate that 1St 1.3 is novel HMW-GS variants which will be valuable for enhancing our understanding of structural differentiation and the evolutionary relationship among HMW-GSs in Triticeae species.     

Isolation and characterization of novel Glu-St1 alleles from Pseudoroegneria spicata and Pd. strigosa

Pseudoroegneria is a small genus of the Triticeae tribe; its St genome is present in over half of allopolyploid Triticeae species. The high molecular weight (HMW) subunits of glutenin (GS) encoded by the St genome are not well described. In this paper, we report the characterization of fourteen alleles of HMW-GS genes from the two species Pd. spicata and Pd. strigosa. Analysis shows that all fourteen sequences possess a typical primary structure shared by other known HMW-GS, but with some unique modifications. All fourteen Glu-St1 alleles are significantly smaller than normal Glu-1 genes due to fewer repeat motifs in a repetitive region with no indication of large deletion in other conserved regions. Thus, the small size is a common feature of HMW-GS encoded by Glu-St1 loci of Pseudoroegneria species. Sequence analysis indicated that all fourteen Glu-St1 alleles were intermediate type between x- and y-type, which represent an intermediate stage in the evolutionary divergence of x- and y-type subunits.     

Molecular cloning, heterologous expression, and phylogenetic analysis of a novel y-type HMW glutenin subunit gene from the G genome of Triticum timopheevi.

A novel y-type high molecular weight (HMW) glutenin subunit gene from the G genome of Triticum timopheevi (2n=4x=28, AAGG) was isolated and characterized. Genomic DNA from accession CWI17006 was amplified and a 2200 bp fragment was obtained. Sequence analysis revealed a complete open reading frame including N- and C-terminal ends and a central repetitive domain encoding 565 amino acid residues. The molecular weight of the deduced subunit was 77,031, close to that of the x-type glutenin subunits. Its mature protein structure, however, demonstrated that it was a typical y-type HMW subunit. To our knowledge, this is the largest y-type subunit gene among Triticum genomes. The molecular structure and phylogenetic analysis assigned it to the G genome and it is the first characterized y-type HMW glutenin subunit gene from T. timopheevi. Comparative analysis and secondary structure prediction showed that the subunit possessed some unique characters, especially 2 large insertions of 45 (6 hexapeptides and a nonapeptide) and 12 (2 hexapeptides) amino acid residues that mainly contributed to its higher molecular weight and allowed more coils to be formed in its tertiary structure. Additionally, more alpha-helixes in the repeat domain of the subunit were found when compared with 3 other y-type subunits. We speculate that these structural characteristics improve the formation of gluten polymer. The novel subunit, expressed as a fusion protein in E. coli, moved more slowly in SDS-PAGE than the subunit Bx7, so it was designated Gy7*. As indicated in previous studies, increased size and more numerous coils and alpha-helixes of the repetitive domain might enhance the functional properties of HMW glutenins. Consequently, the novel Gy7* gene could have greater potential for improving wheat quality.     

PCR analysis of x- and y-type genes present at the complex Glu-A1 locus in durum and bread wheat

Genes (x-type) corresponding to different high-molecular-weight glutenin subunits encoded at the Glu-A1 locus present in bread- and durum-wheat cultivars have been selectively amplified by the polymerase chain reaction (PCR). DNA fragments corresponding to an unexpressed x-type gene were also amplified. As unexpressed y-type genes may or may not contain an 8-kb transposon-like insertion, two different sets of primers were designed to obtain amplification of DNA fragments corresponding to these genes. Amplified DNA fragments were also digested with restriction enzymes. The digestion patterns of amplified fragments corresponding to unusual x-type subunits showed similarities with genes encoding the most common subunits 2^* and 1. The unexpressed amplified x-type gene showed a restriction pattern similar to the one obtained with the allelic gene encoding high-molecular-weight glutenin subunit 1; homologies were also found within the repetitive region of the linked y-type genes. On the basis of these observations it is postulated that an ancestral active x-type gene, most likely corresponding to subunit 1, was silenced following the insertion of the 8-kb transposon-like fragment into the linked y-type gene. Received: 8 April 1996 / Accepted: 30 August 1996     

Characterization of the HMW glutenin subunits from Aegilops searsii and identification of a novel variant HMW glutenin subunit

High molecular weight (HMW) glutenin subunits are conserved seed storage proteins in wheat and related species. Here we describe a more detailed characterization of the HMW glutenin subunits from Aegilops searsii, which is diploid and contains the S^s genome related to the S genome of Aegilops speltoides and the A, B and D genomes of hexaploid wheat. SDS-PAGE experiments revealed two subunits (one x and one y) for each of the nine Ae. searsii accessions analyzed, indicating that the HMW glutenin subunit gene locus of Ae. searsii is similar to the Glu-1 locus found in wheat in containing both x and y genes. The primary structure of the four molecularly cloned subunits (from two Ae. searsii accessions) was highly similar to that of the previously reported x and y subunits. However, in one accession (IG49077), the last 159 residues of the x subunit (1S^sx49077), which contained the sequence element GHCPTSPQQ, were identical to those of the y subunit (1S^sy49077) from the same accession. Consequently, 1S^sx49077 contains an extra cysteine residue located at the C-terminal part of its repetitive domain, which is novel compared to the x-type subunits reported so far. Based on this and previous studies, the structure and expression of the Glu-1 locus in Ae. searsii is discussed. A hypothesis on the genetic mechanism generating the coding sequence for the novel 1S^sx49077 subunit is presented.     

High-Molecular-Weight Glutenin Subunit Genesin in Decaploid Agropyron elongatum

Seven genes encoding glutenin subunits that present in Agropyron elongatum (Host) Nevski were cloned by PCR analysis and named AgeloG1 to AgeloG7. The complete open reading frames (ORFs) of the seven genes were amplified with primers special for high-molecular-weight (HMW) glutenin subunit genes and subsequently cloned and sequenced. Five of them were completely sequenced, and the other two (AgeloG1 and AgeloG4) were sequenced at the two ends only. Comparison of amino acid sequences suggested that the primary structure of the subunits encoded by the seven genes was very similar to that of y-type HMW glutenin subunits published from wheat, though four of them (AgeloG4, AgeloG5, AgeloG6 and AgeloG7) were shorter than 1.8 kb. Phylogenetic analysis of the five completely sequenced genes and those subunit genes of Triticum aestivum L. (AABBDD), Aegilops tauschii Coss. (DD), Aegilops caudata L. (CC), Secale cereale L. (RR) and Aegilops umbellulata Zhuk. (UU) indicated that the AgeloG2 was most closely related to 1Dy; the AgeloG3 was to 1By; the AgeloG5, AgeloG6 and AgeloG7 were to 1Ay.     

小麦高分子量谷蛋白亚基Glu-B1位点沉默基因的克隆与序列分析

二粒小麦（Triticum turgidum L．var．dicoccoides）具有极其丰富的遗传多样性,是栽培小麦品种改良的巨大基因库。在高分子量谷蛋白基因的组成上,它具有许多栽培小麦不存在的变异类型,在Glu—B1位点上的变异更大。我们利用种子贮藏蛋白的SDS—PAGE方法从原产于伊朗的二粒小麦材料PI94640中观察到缺失Glu—B1区的高分子量谷蛋白亚基。利用Glu-1Bx基因保守序列设计PCR引物,对该材料的总DNA扩增,获得了X型亚基编码基因（Glu-1Bxm）的全序列,其全长为3442bp含1070bp的启动子区。序列比较发现,Glu-1Bxm在启动子区序列与Glu—1Bx7的最为相似。而在基因编码区,我们发现Glu—1Bxm仅编码212个氨基酸,由于开放阅读框中起始密码子后第637位核苷酸发生了点突变,即编码谷酰胺的CAA突变为终止密码TAA,可能直接导致了该高分子量谷蛋白亚基的失活,这是我们在小麦Glu—B1位点基因沉默分子证据的首次报道。将Glu—1Bxm全序列与Glu—B1位点其他等位基因进行了系统树分析,发现Glu—1Bxm是较为古老的类型。本文还对该特异高分子量谷蛋白亚基变异类型对品质遗传改良研究的意义进行了讨论。     

Intermolecular disulfide bonds link specific high-molecular-weight glutenin subunits in wheat endosperm.

A detergent wash extracted soluble proteins from wheat flour, leaving a residue enriched with insoluble glutenin aggregates. Digestion of this residue with endoproteinase Lys-C, which showed a limited specificity for glutenin subunits, produced several peptides with apparent molecular weights close to those of intact high-molecular-weight glutenin subunits. N-terminal sequencing indicated that the isolated peptides were composed of high-molecular-weight glutenin subunit fragments joined by an intermolecular disulfide bond. In two of these peptides, only two components were found, one from an x-type subunit and the other from a y-type subunit. The isolated peptides all contained at least one x-type C-terminal region and one y-type N-terminal region, suggesting a specific orientation to the intermolecular disulfide linkage.     

Molecular characterization and phylogenetic analysis of a novel glutenin gene (Dy10.1t) from Aegilops tauschii.

A novel y-type high molecular mass glutenin subunit (HMM-GS) possessing a mobility that is slightly slower than that of the subunit Dy10 obtained by SDS-PAGE, named Dy10.1t, in the wild wheat Aegilops tauschii was identified by 1- and 2-dimensional gel electrophoresis, capillary electrophoresis, and matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS). The gene encoding the HMM subunit Dy10.1t was amplified with allele-specific PCR primers, and the amplified products were cloned and sequenced. The coding domain of the Dy10.1t subunit gene consisted of 1980 bp encoding a protein of 658 residues with an M rs of 68 611 Da, which was similar to the M rs determined by MALDI-TOF-MS. The deduced amino acid sequence indicated that Dy10.1t subunit displayed a greater similarity to the Dy12 subunit, differing by only 8 amino acid substitutions. Six coding region single-nucleotide polymorphisms were discovered in the Dy10.1t gene by multiple alignments (1 per 330 bp), 1 in the N-terminal domain and the others in the central repeats. Five of them resulted in residue substitutions, whereas 3 created enzyme site changes. The homology and neighbour-joining trees constructed from code domain sequences of 20 x- and y-type glutenin genes from different Triticum species separated into 2 halves, which corresponded to the x-type and y-type HMM glutenin alleles. Phylogenetic analysis revealed that the Glu-1 gene duplication event probably occurred at about 16.83 million years ago, whereas the divergence times of A, B, and D genomes within x-type and y-type halves were before 7.047 and 10.54 million years ago, respectively.     

Characterisation of two gene subunits on the 1R chromosome of rye as orthologs of each of the Glu-1 genes of hexaploid wheat

The visco-elastic properties of bread flour are firmly associated with the presence or absence of certain HMW subunits coded by the Glu-1 genes. Identifying allelic specific molecular markers (AS-PCR) associated with the presence of Glu-1 genes can serve as a valuable tool for the selection of useful genotypes. This paper reports the use of primers designed from nucleotide sequences of the Glu-D1 gene of wheat (AS-PCR for Glu-D1y10) that recognise and amplify homologous sequences of the Glu-R1 gene subunits of rye. The primers amplify the complete coding regions and provided two products of different size in rye, in wheats carrying the substitution 1R(1D) and in rye-wheat aneuploid lines carrying the long arm of chromosome 1R. The location, the molecular characterisation of these sequences and their expression during grain ripening seem to demonstrate that the amplification products correspond to structural genes encoding the high-molecular-weight (HMW) glutenins of rye. The homology of the rye gene to subunits encoding HMW glutenins in wheat was confirmed by Southern blots and sequencing. The amplification-products were cloned, sequenced and characterised, and the sequences compared with the main glutenin subunits of wheat and related species. Further, an RT-PCR experiment was performed using primers designed from the sequence of both amplified products. This assay demonstrated that both sequences are expressed in endosperm during grain ripening. The results of these analyses suggest that both gene subunits correspond to x- and y-type genes of the Glu-R1 locus of rye. Received: 11 December 2000 / Accepted: 17 April 2001     

Characterizing HMW-GS alleles of decaploid <Emphasis Type="Italic">Agropyron elongatum</Emphasis> in relation to evolution and wheat breeding

Bread wheat quality is mainly correlated with high molecular weight glutenin subunits (HMW-GS) of endosperm. The number of HMW-GS alleles with good processing quality is limited in bread wheat cultivars, while there are plenty of HMW-GS alleles in wheat-related grasses to exploit. We report here on the cloning and characterization of HMW-GS alleles from the decaploid Agropyron elongatum. Eleven novel HMW-GS alleles were cloned from the grass. Of them, five are x-type and six y-type glutenin subunit genes. Three alleles Aex4, Aey7, and Aey9 showed high similarity with another three alleles from the diploid Lophopyrum elongatum, which provided direct evidence for the E^e genome origination of A. elongatum. It was noted that C-terminal regions of three alleles of the y-type genes Aey8, Aey9, and Aey10 showed more similarity with x-type genes than with other y-type genes. This demonstrates that there is a kind of intermediate state that appeared in the divergence between x- and y-type genes in the HMW-GS evolution. One x-type subunit, Aex4, with an additional cysteine residue, was speculated to be correlated with the good processing quality of wheat introgression lines. Aey4 was deduced to be a chimeric gene from the recombination between another two genes. How the HMW-GS genes of A. elongatum may contribute to the improvement of wheat processing quality are discussed.     

高冰草中高分子量麦谷蛋白亚基的编码基因

通过SDS-PAGE法分析了高冰草(Agropyron elongatum (Host) Nevski)种子麦谷蛋白亚基,发现高冰草的麦谷蛋白亚基种类比普通小麦更加丰富。通过基因组PCR法用高分子量麦谷蛋白亚基基因的特异引物从高冰草核基因组中分离出了7条麦谷蛋白亚基的全编码序列,分别命名为AgeloG1~AgeloG7。其中的5条已进行全序列测定,对AgeloG1和AgeloG4进行了末端测序。尽管其中的4条基因的编码序列(AgeloG4, AgeloG5, AgeloG6和AgeloG7)小于1.8 kb,但是对从克隆到的序列推导出的氨基酸序列与已经发表的小麦高分子量麦谷蛋白亚基序列进行对比分析发现,这些亚基与来自小麦的高分子量麦谷蛋白亚基具有很高的同源性。并且对信号肽、N-、C-末端的氨基酸序列分析显示,这7条序列编码的亚基皆为y-型亚基。用5条全部测序的编码序列与普通小麦的A、B、D、粗山羊草的D、圆柱山羊草的C、伞穗山羊草的U、黑麦的R染色体的编码高分子量麦谷蛋白的序列进行了聚类分析。表明,AgeloG2与小麦1Dy, AgeloG3与小麦1By, AgeloG5、AgeloG6和AgeloG7与小麦1Ay在起源和进化上有较高的相似性。