2-Methylketones are involved in plant defense and fragrance and have industrial applications as flavor additives and for biofuel production. We isolated three genes from the crop plant Solanum melongena (eggplant) and investigated these as candidates for methylketone production. The wild tomato methylketone synthase 2 (ShMKS2), which hydrolyzes β-ketoacyl-acyl carrier proteins (ACP) to release β-ketoacids in the penultimate step of methylketone synthesis, was used as a query to identify three homologs from S. melongena: SmMKS2-1, SmMKS2-2, and SmMKS2-3. Expression and functional characterization of SmMKS2s in E. coli showed that SmMKS2-1 and SmMKS2-2 exhibited the thioesterase activity against different β-ketoacyl-ACP substrates to generate the corresponding saturated and unsaturated β-ketoacids, which can undergo decarboxylation to form their respective 2-methylketone products, whereas SmMKS2-3 showed no activity. SmMKS2-1 was expressed at high level in leaves, stems, roots, flowers, and fruits, whereas expression of SmMKS2-2 and SmMKS2-3 was mainly in flowers and fruits, respectively. Expression of SmMKS2-1 was induced in leaves by mechanical wounding, and by methyl jasmonate or methyl salicylate, but SmMKS2-2 and SmMKS2-3 genes were not induced. SmMKS2-1 is a candidate for methylketone-based defense in eggplant, and both SmMKS2-1 and SmMKS2-2 are novel MKS2 enzymes for biosynthesis of methylketones as feedstocks to biofuel production.
We report here the biosynthesis of daidzein in Streptomyces sp. SS52, its genome sequence and the analysis of its genome for finding putative genes involved in daidzein biosynthesis. The Streptomyces sp. SS52 strain was isolated from the plant Phyllanthus urinaria in Tra Vinh province, Vietnam. This endophytic strain is capable of producing the isoflavone daidzein in the culture medium. Streptomyces sp. SS52 possesses a linear genome of 8,184,045 bp and the GC content of this genome is 72.5%. The preliminary genome analysis identified homologs of genes involved in the de novo biosynthesis of daidzein in the genome of Streptomyces sp. SS52. The genome sequencing of Streptomyces sp. SS52 was essential for the study of the biosynthesis of daidzein in Streptomyces bacteria.
In our previous publication, we have tentatively concluded that our fungal specimen DL0038A and DL0038B are Cordyceps takaomontana. In order to further support the identification, we continued to analyse the nrSSU (nuclear ribosomal small subunit) and rpb1 (largest subunit of RNA polymerase II) genes, as well as combined analysis the nrLSU (largest subunit of RNA polymerase II) together with nrSSU and rpb2 genes on these specimens in order to support the morphological identification of those fungi. The results show that we successfully amplified all genes. Sequencing method was then adopted and proofread before molecular phylogenetic analysis was applied with reference sequences obtained from the publication of Sung et al. (2007). Once again, this analysis strongly supports the DL0038A and DL0038B specimen as Cordyceps takaomontana.
Trichoderma species are known as potential biocontrol agents against the plant pathogenic fungi with various mechanisms (parasitism, antibiosis, substrate competition...). One of the most important mechanisms is the secretion of cell wall degrading enzymes. Chitinolytic enzymes, especially, endochitinase plays an important role in the enzymatic mechanism by the abundant and frequent induction more than other groups in whole of chitinolytic system. Trichoderma longibrachiatum TÐ16 was grown in the TSM medium containing various substrates. We recognized that 1.0% (w/v) colloidal chitin and 0.5% (w/v) S. rolfsii cell wall induced to biosynthesize chitinase and endochitnase with higher activity than P. capsici cell wall at the fifth day. This strain induced and expressed various endochitinase isozymes when growing on the substrates above, respectively. Two endochitnase isozymes (52 and 42 kDa) on colloidal chitin and (42 and 36 kDa) on S. roflsii cell wall were induced. The 42 kDa endochitinase is induced and expressed on both two substrates.
The entomopathogenic fungus T011, parasitizing on nymph of Cicada, collected in the coffee garden in Dak Lak Province, Vietnam, was preliminarily morphologically identified as Isaria cicadae, belonged to order Hypocreales and family Clavicipitaceae. To ensure the authenticity of T011, phylogenetic analysis of the concatenated set of multiple genes including ITS, nrLSU, nrSSU, Rpb1, and Tef1 was applied to support the identification. Genomic DNA was isolated from dried sample T011. The PCR assay sequencing was applied to amplify ITS, nrLSU, nrSSU, Rpb1, and Tef1 gene. For phylogenetic analysis, the concatenated data of both target gens were constructed with MEGAX with a 1,000 replicate bootstrap based on the neighbor-joining, maximum likelihood, maximum parsimony method. As the result, the concatenated data containing 62 sequences belonged to order Hypocreales, families Clavicipitaceae, and 2 outgroup sequences belonged to order Hypocreales, genus Verticillium. The phylogenetic analysis results indicated that T011 was accepted at subclade Cordyceps and significantly formed the monophyletic group with referent Cordyceps cicadae (Telemorph of Isaria cicadae) with high bootstrap value. The phylogenetically analyzed result was strongly supported by our morphological analysis described as the Isaria cicadae. In summary, phylogenetic analyses based on the concatenated dataset were successfully applied to strengthen the identification of T011 as Isaria cicadae.
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