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1.
The caffeic acid O-methyltransferase (COMT) gene is a gene candidate forimproving silage maize digestibility. Before assessing possible functionalconsequences of COMT gene polymorphism, the nucleotide sequence variability atthe COMT locus in 6 normal maize inbred lines was characterized and analyzed.Inaddition, the COMT bm3-3 mutation characterization was specified. Even if theCOMT gene seemed to evolve according to the predictions of the neutral model,estimates of nucleotide diversity indicated that polymorphism distributionwithin the sequence was not uniform and that nonsynonymous sites were the leastvariable. Linkage disequilibrium calculation and estimates of recombinationshowed that polymorphism was a dimorphic segregating distribution caused by alot of recombination events.  相似文献   

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Isolated lignins from alfalfa deficient in caffeic acid 3-O-methyltransferase contained benzodioxanes resulting from the incorporation of the novel monomer, 5-hydroxyconiferyl alcohol. Due to the high level incorporated into the soluble lignin fraction and the use of sensitive NMR instrumentation, unique structural features were revealed. A new type of end-unit, the 5-hydroxyguaiacyl glycerol unit, was identified. It was possible to establish that coniferyl alcohol, sinapyl alcohol, and the novel 5-hydroxyconiferyl alcohol can cross-couple with the 5-hydroxyguaiacyl units that are formed in the lignin, the latter giving rise to extended chains of benzodioxane units. There is also evidence that 5-hydroxyconiferyl alcohol couples with normal (guaiacyl or syringyl) lignin units. Lignin in the alfalfa deficient in caffeoyl CoA 3-O-methyltransferase was structurally similar to the control lignin but the transgenic exhibited a dramatic decrease in lignin content (approximately 20%) and modest increase in cellulose (approximately 10%) reflecting a 30% increase in cellulose:lignin ratio. The compositional changes in both transgenics potentially allow enhanced utilization of alfalfa as a major forage crop by increasing the digestibility of its stem fraction.  相似文献   

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Lignin is a major cell wall component of vascular plants that provides mechanical strength and hydrophobicity to vascular vessels. However, the presence of lignin limits the effective use of crop straw in many agroindustrial processes. Here, we generated transgenic maize plants in which the expression of a lignin biosynthetic gene encoding CCoAOMT, a key enzyme involved in the lignin biosynthesis pathway was downregulated by RNA interference (RNAi). RNAi of CCoAOMT led to significantly downregulated expression of this gene in transgenic maize compared with WT plants. These transgenic plants exhibited a 22.4% decrease in Klason lignin content and a 23.3% increase in cellulose content compared with WT plants, which may reflect compensatory regulation of lignin and cellulose deposition. We also measured the lignin monomer composition of the RNAi plants by GC-MS and determined that transgenic plants had a 57.08% higher S/G ratio than WT plants. In addition, histological staining of lignin with Wiesner reagent produced slightly more coloration in the xylem and sclerenchyma than WT plants. These results provide a foundation for breeding maize with low-lignin content and reveal novel insights about lignin regulation via genetic manipulation of CCoAOMT expression.  相似文献   

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It is generally accepted that peroxidases catalyze the final step in the biosynthesis of lignin. In this study, to examine how expression of prxA3a, a gene for an anionic peroxidase, might be related to lignification in plant tissues, we produced transgenic tobacco plants that harbored a gene for β-glucuronidase (GUS) fused to the prxA3a promoter. Histochemical staining for GUS activity indicated that the prxA3a promoter was active mainly in the lignifying cells of stem tissues. Further, to examine the effects of suppressing the expression of prxA3a, we transferred an antisense prxA3a gene construct into the original host, hybrid aspen (Populus sieboldii ×P. gradidentata), under the control of the original promoter of the prxA3a gene. Eleven transformed aspens were obtained and characterized, and the stable integration of the antisense construct was confirmed by PCR and Southern blotting analysis in all these lines. Assays of enzymatic activity showed that both total peroxidase activity and acidic peroxidase activity were lower in most transgenic lines than in the control plants. In addition, the reduction of peroxidase activity was associated with lower lignin content and modified lignin composition. Transgenic lines with the highest reduction of peroxidase activity displayed a higher syringyl/vanillin (S/V) ratio and a lower S+V yield, mainly because of a decreased amount of V units. Thus, our results indicate that prxA3a is involved in the lignification of xylem tissue and that the down-regulation of anionic peroxidase alters both lignin content and composition in hybrid aspen.  相似文献   

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Lee SH  Hamaker BR 《FEBS letters》2006,580(25):5803-5806
Twenty-seven kilodalton gamma-zein is a subclass of the maize zein storage proteins and, due to its localization at the protein body periphery, is critical to digestibility characteristics of all zeins. This protein had low in vitro digestibility, presumably due to its high Cys content (7.35 mol%) that is similar to the hard-to-digest analogous sorghum protein, gamma-kafirin. Therefore, each of the conserved disulfide-bonded Cys' was mutated to create C144A, C148A, C155A, and C156A maize gamma-zein mutants. The C155A showed a remarkable increase in digestibility to proteases - pepsin, chymotrypsin, and trypsin. A high conservation of this Cys among cereal gamma-prolamins indicates the utility of this finding.  相似文献   

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Cell wall-bound hydroxycinnamic acids and the composition of lignin were studied in relation to the digestibility of a collection of 91 maize silages in wethers. Total lignin and guaiacyl content showed the highest correlation coefficients with digestibility. Using the above-mentioned chemical parameters, eight equations were also developed to predict digestibility. The prediction of organic matter digestibility produced a high adjusted R 2 value (0.487) using total lignin, guaiacyl, esterified ferulic acid and esterified p-coumaric acid content as predictors. The prediction of in vivo dry matter digestibility produced a higher adjusted R 2 value (0.516) using the same variables as predictors. Cell wall digestibility depends on a multiplicity of factors and it is not possible to attribute a causal effect on in vivo digestibility to any single factor. However, total lignin, guaiacyl and p-coumaric acid content emerge as good predictors of digestibility.  相似文献   

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Cell wall-bound hydroxycinnamic acids and the composition of lignin were studied in relation to the digestibility of a collection of 91 maize silages in wethers. Total lignin and guaiacyl content showed the highest correlation coefficients with digestibility. Using the above-mentioned chemical parameters, eight equations were also developed to predict digestibility. The prediction of organic matter digestibility produced a high adjusted R2 value (0.487) using total lignin, guaiacyl, esterified ferulic acid and esterified p-coumaric acid content as predictors. The prediction of in vivo dry matter digestibility produced a higher adjusted R2 value (0.516) using the same variables as predictors. Cell wall digestibility depends on a multiplicity of factors and it is not possible to attribute a causal effect on in vivo digestibility to any single factor. However, total lignin, guaiacyl and p-coumaric acid content emerge as good predictors of digestibility.  相似文献   

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Ma QH  Xu Y 《Biochimie》2008,90(3):515-524
Caffeic acid 3-O-methyltransferase (COMT) catalyzes the multi-step methylation reactions of hydroxylated monomeric lignin precursors, and is believed to occupy a pivotal position in the lignin biosynthetic pathway. A cDNA (TaCM) was identified from wheat and it was found to be expressed constitutively in stem, leaf and root tissues. The deduced amino acid sequence of TaCM showed a high degree of identity with COMT from other plants, particularly in SAM binding motif and the residues responsible for catalytic and substrate specificity. The predicted TaCM three-dimensional structure is very similar with a COMT from alfalfa (MsCOMT), and TaCM protein had high immunoreactive activity with MsCOMT antibody. Kinetic analysis indicated that the recombinant TaCM protein exhibited the highest catalyzing efficiency towards caffeoyl aldehyde and 5-hydroxyconiferaldehyde as substrates, suggesting a pathway leads to S lignin via aldehyde precursors. Authority of TaCM encoding a COMT was confirmed by the expression of antisense TaCM gene in transgenic tobacco which specifically down-regulated the COMT enzyme activity. Lignin analysis showed that the reduction in COMT activity resulted in a marginal decrease in lignin content but sharp reduction in the syringl lignin. Furthermore, the TaCM protein exhibited a strong activity towards ester precursors including caffeoyl-CoA and 5-hydroxyferuloyl-CoA. Our results demonstrate that TaCM is a typical COMT involved in lignin biosynthesis. It also supports the notion, in agreement with a structural analysis, that COMT has a broad substrate preference.  相似文献   

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Caffeic acid O‐methyltransferase (COMT), the lignin biosynthesis gene modified in many brown‐midrib high‐digestibility mutants of maize and sorghum, was targeted for downregulation in the small grain temperate cereal, barley (Hordeum vulgare), to improve straw properties. Phylogenetic and expression analyses identified the barley COMT orthologue(s) expressed in stems, defining a larger gene family than in brachypodium or rice with three COMT genes expressed in lignifying tissues. RNAi significantly reduced stem COMT protein and enzyme activity, and modestly reduced stem lignin content while dramatically changing lignin structure. Lignin syringyl‐to‐guaiacyl ratio was reduced by ~50%, the 5‐hydroxyguaiacyl (5‐OH‐G) unit incorporated into lignin at 10‐–15‐fold higher levels than normal, and the amount of p‐coumaric acid ester‐linked to cell walls was reduced by ~50%. No brown‐midrib phenotype was observed in any RNAi line despite significant COMT suppression and altered lignin. The novel COMT gene family structure in barley highlights the dynamic nature of grass genomes. Redundancy in barley COMTs may explain the absence of brown‐midrib mutants in barley and wheat. The barley COMT RNAi lines nevertheless have the potential to be exploited for bioenergy applications and as animal feed.  相似文献   

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Prediction of organic matter (OM) digestibility (OMD) of primary growth and regrowth grass silages was studied based on their chemical composition, pepsin-cellulase solubility of OM (OMS) and indigestible neutral detergent fibre (INDF) content. Twenty-five primary and 28 regrowth silages were harvested from mixed timothy (Phleum pratense) meadow fescue (Festuca pratensis) or timothy cocksfoot (Dactylis glomerata) swards by varying the dates of the first and second harvest. In vivo OMD was measured with sheep and indigestible NDF was determined by 12 days ruminal incubation of forage samples in nylon bags using dairy cows fed a forage-based diet. Chemical composition of the silages was significantly correlated with digestibility, but single regression equations led to an unacceptable prediction accuracy of OMD (RMSE>40 g/kg DM). Pepsin-cellulase solubility reliably predicted OMD of primary growth silages (RMSE = 10.8 g/kg DM), but was less accurate for regrowth silages (RMSE = 25.9 g/kg). The prediction accuracy of OMD could be improved by using different equations for the two silage types. Indigestible NDF predicted OMD more accurately than OMS for all silages and especially for the regrowth silages. In contrast to OMS, the relationship between INDF and OMD was similar for both silage types.  相似文献   

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The soil organic carbon (SOC) pool is the largest terrestrial reservoir of carbon and plant residues play an important role in its maintenance. Up to 70–80% of SOC in arable soil is composed of humic substances (HS). In these soils post-harvested residues, left in arable soil after harvesting the crops, are the basic source of humus. Previous research indicated that maize plants residue contain a humic acid (HA) fraction possessing recalcitrant compounds that contributed to soil-HA fraction. This study presents updated results obtained using Pyrolysis-Gas Chromatography/Mass Spectrometry (Py-GC/MS) to provide an indication of the contribution of the lignin to the soil HA. Results obtained indicated that the HAs from maize plants were mainly composed of lignin-derived moieties that were likely derived from the partial hydrolysis of p-coumaric and ferulic acid that are linked to lignin, polysaccharides or other biopolymers of the cell wall. Lignin composing the HAs derived from plants and incubated in soil was substantially preserved. Nevertheless the modification of the syringyl/guaiacyl ratio and the oxidation of the side-chains of lignin, suggested a turnover of lignin-derived molecules in soil-HA fraction. This fact indicated an involvement of the alkali insoluble fraction of maize plant residue (humin) in the soil-HA formation, up-dating our previous knowledge.  相似文献   

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Lam TB  Iiyama K  Stone BA 《Phytochemistry》2003,64(2):603-607
The factors affecting in vitro dry matter digestibility (IVDMD) of fully mature internodes of 150 lines of the forage grass, Phalaris aquatica, and internodes of 100 lines of perennial ryegrass (Lolium perenne), harvested just after anthesis, were investigated. The relationships between IVDMD and the contents of acetyl bromide lignin, and ester-ether linkages between lignin and wall polysaccharides, measured by hydroxycinnamic acids (HCAs) released by 4 M NaOH at 170 degrees C respectively, were determined. The regression analysis gave r(2)=0.05 and 0.03 for the relation between IVDMD and lignin content and r(2)=0.51 and 0.53 for the relation between IVDMD and the content of hot alkali-labile HCA (predominantly ferulic acid) for phalaris and ryegrass, respectively. These observations are interpreted in terms of the restricted accessibility of polysaccharide hydrolysing enzymes to their substrates in the forage cell walls by the covalent cross-linking of wall polymers through HCAs.  相似文献   

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Chemical imaging by confocal Raman microscopy has been used for the visualization of the cellulose and lignin distribution in wood cell walls. Lignin reduction in wood can be achieved by, for example, transgenic suppression of a monolignol biosynthesis gene encoding 4-coumarate-CoA ligase (4CL). Here, we use confocal Raman microscopy to compare lignification in wild type and lignin-reduced 4CL transgenic Populus trichocarpa stem wood with spatial resolution that is sub-μm. Analyzing the lignin Raman bands in the spectral region between 1,600 and 1,700 cm−1, differences in lignin signal intensity and localization are mapped in situ. Transgenic reduction of lignin is particularly pronounced in the S2 wall layer of fibers, suggesting that such transgenic approach may help overcome cell wall recalcitrance to wood saccharification. Spatial heterogeneity in the lignin composition, in particular with regard to ethylenic residues, is observed in both samples. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.  相似文献   

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