In this work, analytical pyrolysis (Py-GC-MS) was employed to identify lignin markers derived from H, S, and G phenylpropanoid units in sugarcane bagasse. Temperatures of 450 and 500 • C allowed the detection of key products that were informative on the bagasse lignin composition. The method was validated by comparing the S/G ratio as determined by the nitrobenzene oxidation (NBO) standard method for five sugarcane varieties. The S/G ratio as determined by Py-GC-MS, taking into consideration all known lignin markers in the analysis, resulted in a correlation coefficient of 0.85 with a linear regression coefficient of 0.74. When a group of selected markers (M2) was used, the correlation coefficient between methods was improved to 0.95 and the linear regression coefficient was adjusted to 0.92. M2 markers consisted of five syringyl markers (syringol, 4-methylsyringol, 4-ethylsyringol, 4-vinylsyringol, and trans-4-propenylsyringol) and four guaiacyl markers (guaiacol, 4-vinylguaiacol, 4-methylguaiacol, and vanillin). Importantly, Py-GC-MS allowed for the study of lignin composition in sugarcane bagasse without the need to remove the extractives, minimizing the work with sample preparations.
Penicillium griseoroseum has been studied because of its efficient pectinases production. In this work, the Penicillium griseoroseum nitrate reductase gene was characterized, transcriptionaly analyzed in different nitrogen sources, and used to create a phylogenetic tree and to develop a homologous transformation system. The regulatory region contained consensus signals involved in nitrogen metabolism and the structural region was possibly interrupted by 6 introns coding for a deduced protein with 864 amino acids. RT–PCR analysis revealed high amounts of niaD transcript in the presence of nitrate. Transcription was repressed by ammonium, urea, and glutamine showing an efficient turnover of the niaD mRNA. Phylogenetics analysis showed distinct groups clearly separated in accordance with the classical taxonomy. A mutant with a 122-bp deletion was used in homologous transformation experiments and showed a transformation frequency of 14 transformants/µg DNA. All analyzed transformants showed that both single- and double-crossover recombination occurred at the niaD locus. The establishment of this homologous transformation system is an essential step for the improvement of pectinase production in Penicillium griseoroseum.Key words: nitrate reductase, nitrogen metabolism, Penicillium griseoroseum, phylogenetic analysis, homologous transformation.
Xyloglucan is the major hemicellulosic polymer found in the primary cell walls of dicots. Xyloglucan tethers cellulose microfibrils conferring rigidity and strength for maintenance of cell integrity, and it is thought that its metabolism contributes to cell elongation and thus plant growth. Here, we have cloned and characterized a Eucalyptus grandis gene ortholog of the Arabidopsis thaliana MUR3 gene (xyloglucan galactosyltransferase), thus termed EgMUR3. EgMUR3 represents an intronless sequence of 1,854 bp predicted to encode a protein of 617 amino acid residues. It exhibits 73% identity and 82% similarity to the A. thaliana MUR3 gene. To demonstrate that this gene encodes a functional enzyme, the putative ORF was cloned into a binary vector under the control of a constitutive promoter and transformed into the A. thaliana mur3 mutant. The effect of the genetic complementation was investigated by xyloglucan oligosaccharide fingerprinting of wall material. The results confirmed that EgMUR3 represents indeed a xyloglucan galactosyltransferase of E. grandis able to use endogenous substrate(s) in A. thaliana, suggesting that both species share common steps in xyloglucan biosynthesis.
Sugarcane (Saccharum spp.) bagasse is recognized as a promising feedstock for cellulosic ethanol production. e development of high-quality crop cultivars through plant breeding is a way to improve feedstock quality. is study aimed to directly and indirectly quantify the e ects of agronomic characteristics on the sacchari cation of sugarcane bagasse samples and to identifying the characteristics that can be used as reliable tools for the selection of genotypes for a better biofuel feedstock. e lignin content was determined in bagasse samples of 286 clones from 13 half-sib families. Twenty clones showing a more contrasting lignin content were analyzed for the cellulose and hemicellulose composition in bagasse. ey di ered statistically for cellulose and hemicellulose contents and also for the e ciency of cellulose to glucose conversion. Clones with the highest dry biomass content also had the highest ber content and were grouped among the clones exhibiting the highest lignin content. Lignin content was statistically signi cant and positively correlated with ber (r = 0.485, p < 0.05) and plant biomass (r = 0.654, p < 0.01), and hemicellulose content was statistically signi cant and positively correlated with number of internodes (r = 0.623, p < 0.01). Lignin content and plant biomass had large negative direct e ects (0.405 and 0.784, respectively) on glucose released by enzymatic hydrolysis. Variation among clones for plant productivity parameters and chemical composition of bagasse could be exploited by breeding to increase the amount of cellulosic ethanol produced. Lignin content and plant biomass were the most important components with direct and negative in uences on the conversion of cellulose to glucose.
Background
Water is one of the main limiting factors for plant growth and crop productivity. Plants constantly monitor water availability and can rapidly adjust their metabolism by altering gene expression. This leads to phenotypic plasticity, which aids rapid adaptation to climate changes. Here, we address phenotypic plasticity under drought stress by analyzing differentially expressed genes (DEG) in four phylogenetically related neotropical Bignoniaceae tree species: two from savanna, Handroanthus ochraceus and Tabebuia aurea, and two from seasonally dry tropical forests (SDTF), Handroanthus impetiginosus and Handroanthus serratifolius. To the best of our knowledge, this is the first report of an RNA-Seq study comparing tree species from seasonally dry tropical forest and savanna ecosystems.
Results
Using a completely randomized block design with 4 species × 2 treatments (drought and wet) × 3 blocks (24 plants) and an RNA-seq approach, we detected a higher number of DEGs between treatments for the SDTF species H. serratifolius (3153 up-regulated and 2821 down-regulated under drought) and H. impetiginosus (332 and 207), than for the savanna species. H. ochraceus showed the lowest number of DEGs, with only five up and nine down-regulated genes, while T. aurea exhibited 242 up- and 96 down-regulated genes. The number of shared DEGs among species was not related to habitat of origin or phylogenetic relationship, since both T. aurea and H impetiginosus shared a similar number of DEGs with H. serratifolius. All four species shared a low number of enriched gene ontology (GO) terms and, in general, exhibited different mechanisms of response to water deficit. We also found 175 down-regulated and 255 up-regulated transcription factors from several families, indicating the importance of these master regulators in drought response.
Conclusion
Our findings show that phylogenetically related species may respond differently at gene expression level to drought stress. Savanna species seem to be less responsive to drought at the transcriptional level, likely due to morphological and anatomical adaptations to seasonal drought. The species with the largest geographic range and widest edaphic-climatic niche, H. serratifolius, was the most responsive, exhibiting the highest number of DEG and up- and down-regulated transcription factors (TF).
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