The effect of O‐methyltransferase (OMT) cDNA modulation on cell wall composition, ultrastructure and rumen degradability was measured on transgenic tobacco (Nicotiana tabacum). The expression of OMT cDNA in antisense orientation (AS) inhibited OMT activity by 92% whereas expression of sense constructs led to plants either co‐suppressed (CS, 98% inhibition) or overexpressing OMT activity. The cell wall residues of stems were analysed for lignin content, products of nitrobenzene oxidation (NBO) and polysaccharide content. Degradability was determined by a cellulase method. Sections of stem were stained by acid phloroglucinol and Mäule reactant. Stem samples were incubated in the rumen for 8, 24 and 48 h and observed by scanning electron microscopy (SEM). Compared to controls, OMT‐depleted stems showed decreased hemicellulose content but unchanged lignin content. In contrast, syringyl units decreased by 40 and 90% in AS and CS samples respectively and NBO content followed a similar trend. Dry matter cellulase degradability was significantly improved by 3·5 and 5·6 percentage units in AS and CS samples respectively. SEM showed a greater bacterial colonisation in these samples and indicated a higher rate of rumen degradability in CS tissues than in controls. Overexpressing plants had a composition and a degradability similar to that of controls. For all the plants studied, the improvements in dry matter degradability were closely linked to the syringyl to guaiacyl ratio or to the NBO content. The modifications observed in down‐regulated tobacco were similar to those produced by bm3 maize mutation, but without lignin decrease. Genetic modifications should therefore be considered for improving forage digestibility.
The effect of down‐regulation of tobacco cinnamyl alcohol dehydrogenase (CAD) on cell wall composition and degradability has been assessed. CAD activity was only 20, 16, 14 and 7%, relative to the controls, in four populations of plants (designated 40‐1, 40‐2, 48 and 50, respectively) transformed with CAD antisense mRNA. Cell wall residues of stem samples were analysed for polysaccharide composition, gravimetric and acetyl bromide lignins and lignin nitrobenzene oxidation products. In situ disappearance and cellulase solubility of both initial dry matter and CWR were determined. The populations of plants with depressed CAD activity showed no change in lignin content but some consistent changes in cell wall composition and digestibility were identified. The syringyl content of lignins decreased and the syringaldehyde to vanillin ratio (S/V) was consequently reduced. Dry matter degradability, as measured by both methods, was significantly improved in all CAD‐depressed samples except for population 40‐1, which was the least CAD‐depressed. Increased in situ disappearance of cell wall (ISCWD) was found in all plants exhibiting more than 80% CAD down‐regulation and was maximal (7 percentage units) in population 50 which had the greatest CAD depression. The rates of ISCWD increased slightly in some populations (40‐2 and 50). The relationship between S/V and ISCWD was significant (R = ‐0·68) only in the samples from a selected population of mature, most depleted plants. Other modifications may therefore also contribute to the improvement in degradability. However the changes in lignin composition that were observed in CAD‐depressed tobacco are largely similar to those seen in some maize and sorghum mutants with altered lignification and improved digestibility. These data therefore suggest that depressing CAD activity may be an effective method for improving digestibility in forage crops. © 1998 SCI.
Apical internodes of tall fescue (Festuca arundinacea Schreb. var. Clarine) harvested at flowering were sectioned into 5 or 10 equal parts to study in situ degradability and cell wall composition, respectively. The basal (youngest) section had the greatest primary wall content. Cell walls in the upper (older) sections had the highest xylose/arabinose ratio and lignin content and a lignin rich in syringyl units, all typical of extensive secondary wall development. Almost all of the p-coumaric (p-CA) and about half of the ferulic acid (FA) were released by 1 M NaOH and presumed to be ester-linked. The total FA content was approximately double that of p-CA in all sections other than the youngest with a distribution similar to that of total p-CA. However, the ratio of esterified to ether and ether plus ester linked (Et & Et+Es) FA differed with age. Whereas the esterified form remained essentially constant ( approximately 4.5 g/kg of cell wall), Et & Et+Es ferulate increased with increasing age of the tissue and was significantly related to lignin deposition (r = 0.79, P < 0.01). The extent of cell wall degradation after 48 h of incubation in the rumen was inversely related to maturity, falling from 835 g/kg of dry matter in the youngest section to 396 g/kg in the oldest. Both the rate and extent of cell wall degradation were significantly negatively related to the ratio of xylose to arabinose, lignin content, proportion of syringyl units present in lignin, and concentration of Et & Et+Es FA present. A positive relationship between Et & Et+Es FA was also found, with the rate (P < 0.01) being better correlated than the extent (P < 0.05) of cell wall degradation. Application of the newly extended internode model to fescue produced results consistent with the view that both the lignin content and the extent to which lignin was covalently bound to the other wall polymers crucially influenced the rate and extent of degradation.
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