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根据小麦低分子量谷蛋白基因保守区序列设计引物P1/P2, 采用PCR法对四川小麦地方品种AS1643的基因组DNA进行扩增, 获得1条约900 bp的片段, 分离、纯化后连接到载体pMD18-T上, 对筛选阳性克隆测序, 获得1个低分子量谷蛋白基因LMW-AS1643(GenBank登录号: EF190322), 其编码区长度为909 bp, 可编码302个氨基酸残基组成的成熟蛋白。序列分析结果表明, LMW-AS1643具有典型的低分子量谷蛋白基因的基本结构, 其推导氨基酸序列与其它已知的LMW-GS相比, 最高相似性为93.40%。生物信息学分析表明, 在LMW-AS1643低分子量谷蛋白中, 无规则卷曲含量最高, 为67.90 %, 其次是a-螺旋, 占30.46 %, b-折叠含量最少, 为1.64 %。 相似文献
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Allelic variation of the low-molecular-weight glutenin subunit (LMW-GS) is associated with the significant differences of dough quality in bread and durum wheat, and has been widely evaluated at protein level in wheat and its relatives. In this study, a PCR primer set, targeting the high variable repetitive domains, was employed to assay the length variation of i-type LMW-GS genes in the A-genomes of diploid wheats, the diploid progenitors of tetraploid and hexaploid wheat. A total of 71 accessions of diploid wheats, belonging to two wild and one cultivated species, were investigated. The higher variations of repetitive length in i-type LMW-GS genes were found in diploid wheats with Nei's genetic variation index (H) of 0.834. The two wild species, T. boeoticum and T. urartu, were found to possess the similar degree of variability, with the Nei's genetic variation index of 0.806 and 0.783, respectively. Less variations were detected in T. monococcum (H = 0.680), a cultivated species domesticated from T. boeoticum. The sufficient variations found in this study could be used as valuable sources for the enrichment of the genetic variations and the alteration of flour-processing properties of the cultivated wheat. To our knowledge, it was the first time that an analysis of length variation targeting a particular group of genes of LMW-GS complex multigene families was conducted. 相似文献
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H. Long Z. Huang Y. -M. Wei Z. -H. Yan Z. -C. Ma Y. -L. Zheng 《Russian Journal of Genetics》2008,44(4):429-435
Allelic variation of the low-molecular-weight glutenin subunit (LMW-GS) is associated with the significant differences of
dough quality in bread and durum wheat, and has been widely evaluated at protein level in wheat and its relatives. In this
study, a PCR primer set, targeting the high variable repetitive domains, was employed to assay the length variation of i-type
LMW-GS genes in the A-genomes of diploid wheats, the diploid progenitors of tetraploid and hexaploid wheat. A total of 71
accessions of diploid wheats, belonging to two wild and one cultivated species, were investigated. The higher variations of
repetitive length in i-type LMW-GS genes were found in diploid wheats with Nei’s genetic variation index (H) of 0.834. The two wild species, T. boeoticum and T. urartu, were found to possess the similar degree of variability, with the Nei’s genetic variation index of 0.806 and 0.783, respectively.
Less variation was detected in T. monococcum (H = 0.680), a cultivated species domesticated from T. boeoticum. The sufficient variation found in this study could be used as valuable source for the enrichment of genetic variations and
the alteration of flour-processing properties of the cultivated wheat. To our knowledge, it was the first time that an analysis
of length variation targeting a particular group of genes of LMW-GS complex multigene families was conducted.
This article was submitted by the authors in English. 相似文献
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Ikeda TM Nagamine T Fukuoka H Yano H 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2002,104(4):680-687
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.
Received: 26 March 2001 / Accepted: 16 July 2001 相似文献
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B. G. Cassidy J. Dvorak 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1991,81(5):653-660
Summary A full-length, low-molecular-weight (LMW) glutenin cDNA clone, pTdUCD1, has been isolated from a Triticum durum cv Mexicali wheat cDNA library. The complete sequence was determined and compared to the LMW glutenin genes that have been isolated from hexaploid wheat, Triticum aestivum. This cDNA codes for a protein of 295 amino acids (33,414 daltons) including a 20-amino acid signal peptide as deduced from the DNA sequence. Northern analysis showed that this cDNA hybridizes to a family of related sequences ranging in length from 1,200 to 1,000 nucleotides. This gene is similar but not identical to previously published LMW glutenin gene sequences. The most striking characteristic of all cloned LMW glutenin genes is the conservation of eight cysteine residues, which could be involved in potential secondary or tertiary structure, disulfide bond interactions. This paper presents a structural map defining distinct regions of the LMW glutenin gene family. 相似文献
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Y.-K. Lee M. Ciaffi R. Appels M. K. Morell 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1999,98(1):126-134
Three accessions of T. boeoticum were selected for the cloning and sequencing of novel low-molecular-weight glutenin subunit (LMW-GS) genes, based on the
results of SDS-PAGE and PCR analyses of the LMW-GS diversity in A-genome wheat (Lee et al. 1998 a). A comparison of the nucleotide
and deduced amino-acid sequences of three cloned genes, LMWG-E2, LMWG-E4 and LMWG-AQ1, both to each other and to other known
LMW-GS genes was carried out. The N-terminal domains showed one variable position; GAG (coding for a glutamic acid) for the E-type, and GAT (coding for an aspartic acid) for the Q-type. The comparisons of the LMW-GSs in the literature and this paper define three
different types of N-terminal sequences; METSCIPGLERPW and MDTSCIPGLERPW from the durum and A-genome wheats, and METRCIPGLERPW from the hexaploid and D-genome wheats. The repetitive domains were AC-rich at the nucleotide level and coded for
a large number of glutamine residues; this region showed 16 variable positions changing 12 amino-acid residues, three triple
nucleotide deletions/additions, a large deletion of 18 nucleotides in LMWG-E4 and a deletion of 12 nucleotides in LMWG-E2.
In the C-terminal domains 26 variable positions were found and 12 of these mutations changed amino-acid residues; no deletions/
additions were present in this region. It was shown that the LMWG-E2 and LMWG-E4 genes could be expressed in bacteria and
this allowed the respective protein products to be related back to the proteins defined as LMW-GSs in vivo.
Received: 24 November 1997 / Accepted: 18 August 1998 相似文献
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The thirty-three 5' flanking conserved sequences of the known low-molecular-weight subunit (LMW-GS) genes have been divided into eight clusters, which was in agreement with the classification based on the deduced N-terminal protein sequences. The DNA polymorphism between the eight clusters was obtained by sequence alignment, and a total of 34 polymorphic positions were observed in the approximately 200 bp regions, among which 18 polymorphic positions were candidate SNPs. Seven cluster-specific primer sets were designed for seven out of eight clusters containing cluster-specific bases, with which the genomic DNA of the ditelosomic lines of group 1 chromosomes of a wheat variety 'Chinese Spring' was employed to carry out chromosome assignment. The subsequent cloning and DNA sequencing of PCR fragments validated the sequences specificity of the 5' flanking conserved sequences between LMW-GS gene groups in different genomes. These results suggested that the coding and 5' flanking regions of LMW-GS genes are likely to have evolved in a concerted fashion. The seven primer sets developed in this study could be used to isolate the complete ORFs of seven groups of LMW-GS genes, respectively, and therefore possess great value for further research in the contributions of a single LMW-GS gene to wheat quality in the complex genetic background and the efficient selections of quality-related components in breeding programs. 相似文献
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McIntire TM Lew EJ Adalsteins AE Blechl A Anderson OD Brant DA Kasarda DD 《Biopolymers》2005,78(2):53-61
The high-molecular-weight glutenin subunits (HMW-GS) of wheat gluten in their native form are incorporated into an intermolecularly disulfide-linked, polymeric system that gives rise to the elasticity of wheat flour doughs. These protein subunits range in molecular weight from about 70 K-90 K and are made up of small N-terminal and C-terminal domains and a large central domain that consists of repeating sequences rich in glutamine, proline, and glycine. The cysteines involved in forming intra- and intermolecular disulfide bonds are found in, or close to, the N- and C-terminal domains. A model has been proposed in which the repeating sequence domain of the HMW-GS forms a rod-like beta-spiral with length near 50 nm and diameter near 2 nm. We have sought to examine this model by using noncontact atomic force microscopy (NCAFM) to image a hybrid HMW-GS in which the N-terminal domain of subunit Dy10 has replaced the N-terminal domain of subunit Dx5. This hybrid subunit, coded by a transgene overexpressed in transgenic wheat, has the unusual characteristic of forming, in vivo, not only polymeric forms, but also a monomer in which a single disulfide bond links the C-terminal domain to the N-terminal domain, replacing the two intermolecular disulfide bonds normally formed by the corresponding cysteine side chains. No such monomeric subunits have been observed in normal wheat lines, only polymeric forms. NCAFM of the native, unreduced 93 K monomer showed fibrils of varying lengths but a length of about 110 nm was particularly noticeable whereas the reduced form showed rod-like structures with a length of about 300 nm or greater. The 110 nm fibrils may represent the length of the disulfide-linked monomer, in which case they would not be in accord with the beta-spiral model, but would favor a more extended conformation for the polypeptide chain, possibly polyproline II. 相似文献
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Three novel low-molecular-weight glutenin subunit (LMW-GS) genes (designated as Ht1, Ht2, and Ht3) were isolated from the genomic DNA of Hordeum brevisubulatum ssp. turkestanicum by PCR amplification (accession no. Y0695). The coding regions of Ht1, Ht2, and Ht3 were 924, 924, and 903 bp, respectively. The deduced amino acid sequences were 306, 306, and 299 amino acid residues each
with a signal peptide, a central repetitive region rich in proline and glutamine, and N-and C-terminal non-repetitive domains.
A comparison was carried out of these genes with other known B hordein genes from cultivated barley and LMW glutenin genes
from wheat. The results indicated that Ht1, Ht2, and Ht3 had a more similar structure and a higher level of homology with the LMW-GS genes than the B hordein genes. In order to investigate
the evolutionary relationship of the novel genes with the prolamin genes from barley and wheat, the phylogenetic tree was
constructed and the subfamilies of these prolamin genes were identified. The results suggested that the three novel genes
were glutenin-like proteins designated as LMW-m type genes.
The text was submitted by the authors in English. 相似文献
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M. Ciaffi Y. K. Lee L. Tamas R. Gupta J. Skerritt R. Appels 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1999,98(1):135-148
The isolation and characterisation by DNA sequencing of two different low molecular weight glutenin subunit (LMW-GS) genes
from a genomic library derived from Triticum tauschii is described. These genes are similar (more than 90% similarity) but not identical to previously published LMW-GS gene sequences
from cultivated wheats. A comparison of nucleotide sequence of the coding regions revealed the presence of insertions and
deletions preferentially located in the region encoding the domains in the LMW-GS proteins rich in proline and glutamine and
the middle part of the C-domain. The signal sequences, the amino-terminus and the remaining parts of the C-domain were conserved
between all the LMW-GSs compared. The differences detected between the deduced amino-acid sequences in these three regions
are only due to single nucleotide substitutions. The most important characteristic of all compared LMW-GS genes is the conservation
of eight cysteine residues that could be involved in potential secondary or tertiary structure and disulphide-bond interactions.
Comparisons between the 5′ and 3′ non-coding sequences of one of the isolated clones (LMW-16/10) with those of different prolamin
genes from wheat, barley and rye led to the distinction of five different gene families, and confirmed the evolutionary relationships
determined previously for these genes mainly on the basis of the coding region. In particular, the LMW-GS sequences are more
closely related to the B-hordein sequences than to any other prolamin genes from wheat, barley and rye. Formal proof that
the isolated genes coded for LMW-GSs, as defined by gel electrophoresis, was obtained by moving one of these genes (LMW-16/10)
into a bacterial expression vector based on bacteriophage T7 RNA polymerase. The resulting plasmid directed the synthesis
of large amounts of the mature form of the subunit in Escherichia coli. This protein exhibited solubility characteristics identical to those of the LMW-GSs and cross-reacted with antibodies reactive
with these proteins.
Received: 24 November 1997 / Accepted: 18 August 1998 相似文献
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Variation and classification of B low-molecular-weight glutenin subunit alleles in durum wheat 总被引:7,自引:0,他引:7
M. T. Nieto-Taladriz M. Ruiz M. C. Martínez J. F. Vázquez J. M. Carrillo 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》1997,95(7):1155-1160
The B low-molecular-weight (LMW) glutenin subunit composition of a collection of 88 durum wheat cultivars was analyzed. Extensive
variation has been found and 18 different patterns were detected. Each cultivar exhibited 4–8 subunits, and altogether 20
subunits of different mobility were identified. The genetic control of all these subunits was determined through the analysis
of nine F2 populations and one backcross. Five subunits were controlled at the Glu-A3 locus, 14 at Glu-B3 and 1 at Glu-B2. At the Glu-A3 locus each cultivar possessed from zero to three bands and eight alleles were identified. At the Glu-B3 locus each cultivar showed four or five bands and nine alleles were detected. Only one band was encoded by the Glu-B2 locus. A nomenclature for these alleles is proposed and the relationship between them and the commonly used LMW-model nomenclature
is discussed.
Received: 10 February 1997 / Accepted: 25 April 1997 相似文献
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Production and characterization of a transgenic bread wheat line over-expressing a low-molecular-weight glutenin subunit gene 总被引:5,自引:0,他引:5
Masci S. D'Ovidio R. Scossa F. Patacchini C. Lafiandra D. Anderson O.D. Blechl A.E. 《Molecular breeding : new strategies in plant improvement》2003,12(3):209-222
The end-use properties, and thus the value, of wheat flours are determined to a large extent by the proteins that make up the polymeric network called gluten. Low molecular weight glutenin subunits (LMW-GS) are important components of gluten structure. Their relative amounts and/or the presence of specific components can influence dough visco-elasticity, a property that is correlated with the end-use properties of wheat flour. For these reasons, manipulation of gluten dough strength and elasticity is important. We are pursuing this goal by transforming the bread wheat cultivar Bobwhite with a LMW-GS gene driven by its own promoter. Particle bombardment of immature embryos produced several transgenic lines, one of which over-expressed the LMW-GS transgene. Southern blots confirmed that the transgene was integrated into the wheat genome, although segregation analyses showed that its expression was sometimes poorly transmitted to progeny. We have determined that the transgene-encoded LMW-GS accumulates to very high levels in seeds of this line, and that it is incorporated into the glutenin polymer, nearly doubling its overall amount. However, SDS sedimentation test values were lower from the transgenic material compared to a non transgenic flour. These results suggest that the widely accepted correlation between the amount of the glutenin polymers and flour technological properties might not be valid, depending on the components of the polymer. 相似文献
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Isolation of low-molecular-weight glutenin subunit genes from wild emmer wheat (Triticum dicoccoides) 总被引:2,自引:0,他引:2
Three low-molecular-weight glutenin subunit (LMW-GS) genes, designated LMW-Td1, LMW-Td2 and LMW-Td3, were isolated from wild emmer wheat (Triticum dicoccoides), which is the tetraploid progenitor of common wheat (T. aestivum). The complete nucleotide sequence lengths of LMW-Td1, LMW-Td2 and LMW-Td3 are 858, 900 and 1062 bp, respectively. LMW-Td1 and LMW-Td3 can encode proteins with 284 and 352 amino acid residues, respectively, whereas LMW-Td2 is a putative pseudogene due to the presence of 3 inframe stop codons in its C-terminal domain. The deduced protein sequences of the 3 genes share the same typical polypeptide structures with known LMW-GS genes containing 8 cysteines in the mature protein domains. LMW-Td1 was clearly distinguished from all known LMW-GS genes, and considered as a novel LMW-GS gene. Two hydrophobic motifs (i.e. PIIIL and PVIIL) were observed in the repetitive domain of LMW-Td3. Sequence comparison indicates that sequences of the 3 LMW-GS genes from this study are strongly similar to known LMW-GS genes. Our phylogenetic analysis suggests that LMW-Td1 and LMW-Td2 are homologous with genes on chromosome 1A, and LMW-Td3 is closely related to genes on chromosome 1B. 相似文献
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Genetic polymorphism in low-molecular-weight glutenin genes from Triticum aestivum, variety Chinese Spring 总被引:1,自引:0,他引:1
M. Benmoussa L.-P. Vézina M. Pagé S. Yelle S. Laberge 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2000,100(5):789-793
Low-molecular-weight (LMW) glutenin subunits consist mainly of two domains, one at the N- terminus which contains repeats
of short amino-acid motifs, and a non-repetitive one rich in cysteine, at the C- terminal region. In previous reports, polyacrylamide-gel
electrophoresis has been used to show that large size variation exists among LMW and HMW glutenin subunits, and it has been
suggested that deletions and insertions within the repetitive region are responsible for these variations in length. In this
study, PCR-amplification of genomic DNA (Triticum aestivum variety Chinese Spring) was used to isolate three full-length LMW glutenin genes: LMWG-MB1, LMWG-MB2 and LMWG-MB3. The deduced
amino-acid sequences show a high similarity between these ORFs, and with those of other LMW glutenin genes. Comparisons indicate
that LMWG-MB1 has probably lost a 12-bp fragment through deletion and that LMWG-MB1 and LMWG-MB2 have an insertion of 81 bp
within the repetitive domain. The current study has shown direct evidence that insertions and/or deletions provide a mechanistic
explanation for the allelic variation, and the resultant evolution, of prolamin genes. Single-base substitutions at identical
sites generate stop codons in both LMWG-MB2 and LMWG-MB3 indicating that these clones are pseudogenes.
Received: 7 May 1999 / Accepted: 17 June 1999 相似文献
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Functional properties of a new low-molecular-weight glutenin subunit gene from a bread wheat cultivar 总被引:3,自引:0,他引:3
Xu H Wang RJ Shen X Zhao YL Sun GL Zhao HX Guo AG 《TAG. Theoretical and applied genetics. Theoretische und angewandte Genetik》2006,113(7):1295-1303
Some allelic forms of low-molecular-weight glutenin subunit (LMW-GS) can greatly influence the end-use of wheat flours, understanding the function of each allele of LMW-GS is important to wheat quality breeding. A LMW-GS gene XYGluD3-LMWGS 1(AY263369) has been cloned from bread wheat cultivar Xiaoyan 6. The deduced protein contained nine cystine residues, one more than that in all other LMW-GSs reported previously, indicating that it is either a new gene or a new allele of a known LMW-GS gene. In this study, the gene was expressed in E. coil in large scale for the testing of its functional property. Reactive Red 120-Agarose resin was used efficiently to purify the expressed LMW-GS proteins from bacteria, with the lactic acid–sodium lactate buffer (pH 4.5) which contained low concentration SDS as elution solution. The purified protein (belonging to the LMW-m family, MW about 35 KDa) was supplemented into a base flour, the results of 10 g dough mixing test indicated that incorporation of the LMW-GS increased the strength of the dough, with significant increases in mixing time (MT) and peak width (PW), and decrease in breakdown in resistance (RBD) compared with the control. In addition, the dough with incorporation of the LMW-GS had more glutenin macropolyeric protein than the control, suggesting that the LMW-GS participated in forming larger glutenin polymers, and greatly contributed to dough strength. The changes in mixing parameters and the amount of glutenin macropolyeric protein were related to the quantity of incorporating subunits. 相似文献