Biochemical characterization of a putative wheat caffeic acid O-methyltransferase

A wheat (Triticum aestivum L., near isogenic line of Hamlet) O-methyltransferase (OMT) was previously reported as a putative caffeic acid OMT (TaCOMT1), involved in lignin biosynthesis, based on its high sequence similarity with a number of graminaceous COMTs. The fact that the putative TaCOMT1 exhibits a significantly high sequence homology to another recently characterized wheat flavone-specific OMT (TaOMT2), and that molecular modeling studies indicated several conserved amino acid residues involved in substrate binding and catalysis of both proteins, prompted an investigation of its appropriate substrate specificity. We report here that TaCOMT1 exhibits highest preference for the flavone tricetin, and lowest activity with the lignin precursors, caffeic acid/5-hydroxyferulic acid as the methyl acceptor molecules, indicating that it is not involved in lignin biosynthesis. We recommend its reannotation to a flavone-specific TaOMT1 that is distinct from TaOMT2.     

Metabolic profiles of genetically modified potatoes using a combination of metabolite fingerprinting and multivariate analysis

Comprehensive metabolite fingerprinting of transgenic potatoes that constitutively express human beta amyloid, curdlan synthase (CRDS), and glycogen synthase (glgA); and of wild-type potatoes was carried out using FT-IR and ¹H NMR spectroscopy in combination with multivariate analyses. Comparison of metabolic patterns between transgenic and wild-type potatoes revealed that there were neither quantitative nor qualitative differences in metabolites between transgenic potatoes expressing human beta amyloid, CRDS or glgA, and non-transformed control potatoes. However, there were metabolic differences between two control potato lines — one that was fresh and the other stored. After 1 week of storage, comprehensive metabolite patterns were significantly modified. Although the differences between CRDS and glgA transgenic and control potato lines were small, PCA analysis of FT-IR and ¹H NMR spectral data identified two distinct control lines. These results suggest that the comprehensive metabolite changes in control potato lines, which occurred after 1 week of storage, were greater than the differences between CRDS and glgA transgenic and wild-type potato lines. Thus, the combination of FT-IR and ¹H NMR spectral data and multivariate analysis was valuable for the detection of comprehensive differences in metabolic profiles between transgenic and non-transformed control plants, even though peak-signal overlap prevented assignment of pure compounds. The combination of FT-IR and ¹H NMR spectral data and multivariate analysis is a simple and rapid method for evaluation of the metabolic equivalence of GM crops.     

New retention mechanism of mononucleotides with buffer concentrations in ion-suppressing RP-HPLC

HPLC separation of ionic samples tends to be more complicated and difficult to understand than that of non-ionic compounds. On the other hand, band spacing is much more easily manipulated for ionic than for neutral samples. Ion-suppressing RP-HPLC method was used with organic modifier and aqueous buffer solution. In this work, five mononucleotides of cytidine-5-monophosphate (5′-CMP) disodium salt, uridine-5-monophosphate disodium salt (5′-UMP), guanosine-5-monophosphate disodium salt (5′-GMP), inosine-5-monophosphate disodium salt (5′-IMP), and adenosine-5-monophosphate disodium salt (5′-AMP) were examined. Acetic acid and sodium phosphate were used as buffers, and methanol as an organic modifier. A new relationship between the retention factor and the buffer concentration at a fixed modifier content (5% of methanol) could be expressed by fol|lowing:k=(k ₋₁+k ₀ (K _B/K _S)C _B ^a)/(1+(K _B/K _S)C _B ^a), whereC _B was the concentration of buffer. Using this relationship, the calculated values closely matched the experimental data. The derived relationship showed that as the buffer concentration increased, the retention factor approached a certain value, and this was buffer dependent.     

Isolation and characterization of kelch repeat-containing F-box proteins from colored wheat

Molecular Biology Reports - F-box proteins play important roles in the regulation of various developmental processes in plants. Approximately 1796 F-box genes have been identified in the wheat...     

The phylogeny of fluorescent pseudomonads in an unflooded rice paddy soil

The purpose of this research was to determine the diversity and distribution of fluorescent pseudomonads in an unflooded rice paddy soil. A region of the 16S ribosomal RNA gene from isolates was amplified using PCR and subsequently analysed by sequence analysis for bacterial identification and phylogenetic classification. A total of 117 fluorescent pseudomonads, representing between 10 and 21 species, were isolated from two sampling sites within the same paddy (designated as soils C and S). The isolates were found to be ≥96% homologous with known sequences, and were most closely related to the followingPseudomonas species:P. antarctica, P. costantini, P. extremorientalis, P. frederiksbergensis, P. kilonensis, P. koreensis, P. lini, P. mandelii, P. poae, P. rhodesiae, andP. veronii. Of these matches, the bulk of the isolates (49%) were affiliated withP. mandelii. In soils C and S, phylogenetic analysis revealed that 35 and 82 isolates co-clustered with 39 and 59% of 66 fluorescent pseudomonad type strains, respectively.     

Bacillus thuringiensis serovar mogi (flagellar serotype 3a3b3d), a novel serogroup with a mosquitocidal activity

The flagellated vegetative cells of the Bacillus thuringiensis strain K4 were agglutinated with the H3 reference antiserum and further, agglutinated with 3b and 3d monospecific factor sera but non-reactive for 3c and 3e factor sera. This creates a new serogroup with flagellar antigenic structure of 3a3b3d: B. thuringiensis serovar mogi. The strain K4 showed high activity against dipteran larvae, Anopheles sinensis and Culex pipienspallens while no lepidopteran toxicity. It produced ovoidal parasporal inclusions (crystals) whose SDS-PAGE protein profile consisted of several bands ranging from 75 to 30 kDa. Through the protein identification by nano-LC-ESI-IT MS analysis, the putative peptides of Cry19Ba, Cry40ORF2, Cry27Aa and Cry20Aa were detected.     

Octopamine and 5‐hydroxytryptamine mediate hemocytic phagocytosis and nodule formation via eicosanoids in the beet armyworm,Spodoptera exigua

Octopamine and 5‐hydroxytryptamine (5‐HT) have been known to mediate cellular immune responses, such as hemocytic phagocytosis and nodule formation, during bacterial invasion in some insects. In addition, eicosanoids also mediate these cellular immune reactions in various insects, resulting in clearing the bacteria circulating in the hemolymph. This study investigated a hypothesis on signal cross‐talk between both types of immune mediators in the beet armyworm, Spodoptera exigua, which had been observed in the effect of eicosanoids on mediating the cellular immune responses. In response to bacterial infection, octopamine or 5‐HT markedly enhanced both hemocytic phagocytosis and nodule formation in S. exigua larvae. Their specific antagonists, phentolamine (an octopamine antagonist) or ketanserin (a 5‐HT antagonist) suppressed both cellular immune responses of S. exigua. These effects of biogenic monoamines on the immune mediation were expressed through eicosanoids because the inhibitory effects of both antagonists were rescued by the addition of arachidonic acid (a precursor of eicosanoid biosynthesis). Furthermore, the stimulatory effects of both monoamines on the cellular immune responses were significantly suppressed by different inhibitors acting at their specific levels of eicosanoid biosynthesis. Taken together, this study suggests that octopamine and 5‐HT can mediate hemocytic phagocytosis and nodule formation through a downstream signal pathway relayed by eicosanoids in S. exigua. © 2009 Wiley Periodicals, Inc.     

Fourier transform infrared (FT-IR) spectroscopy of genomic DNA to discriminate F1 progenies from their paternal lineage of Chinese cabbage (Brassica rapa subsp. pekinensis)

Fourier transform infrared spectroscopy (FT-IR) spectroscopy in combination with multivariate analysis was used to discriminate two different F1 hybrid lines from their parental inbred lines. Genomic DNA was isolated from leaves of three parental lines and two F1 hybrid lines of Brassica campestris subsp. pekinensis. Purified genomic DNA was analyzed by FT-IR spectroscopy in the spectral region from 4,000 to 400 cm^?1. FT-IR spectra confirmed typical spectral differences between the frequency regions of N–H stretching (amide I) and C=O stretching vibrations (amide II) as well as PO₂ ^? ionized asymmetric and symmetric stretching. Principal component analysis was able to discriminate between F1 hybrid progenies depending on their parental lineages, even though they share the same maternal background. Partial least squares discriminant analysis gave a more clear discrimination between the two parental lines and their hybrid progenies. These FT-IR spectral differences might be directly related to subtle changes in the base functional group and backbone structures of genomic DNA. Considering these results, this technique could provide a solid research foundation for FT-IR spectral-based rapid diagnosis, selection, and discrimination of parental lines from their progenies. Furthermore, this technique could be applied to test purity in the hybrid seed industry.     

Synechocystis PCC6803 and PCC6906 dnaK2 expression confers salt and oxidative stress tolerance in Arabidopsis via reduction of hydrogen peroxide accumulation

Abiotic stress slows plant growth and development. Because salt stress, particularly from NaCl, acts as an important limiting factor in agricultural productivity, the identification and manipulation of genes related to salt tolerance could improve crop productivity. Prokaryotic, heat shock protein (Hsp), DnaK from the ubiquitous Hsp70 family is upregulated in cells that are under abiotic stress. Synechocystis spp. cyanobacteria encode at least three potential DnaK proteins in their genome. Here, expressions of dnaK1s and dnaK2s from two Synechocystis spp. PCC6803 (Sy6803) and PCC6906 (Sy6906), enhanced salt tolerance in a dnaK-defective Escherichia coli strain. In contrast, dnaK3s in both strains were ineffective, indicating that dnaK3 is functionally different from dnaK1 and dnaK2 in Synechocystis spp. under salt stress. Ectopic expression of dnaK2s from Sy6803 and Sy6906 conferred salt tolerance in transgenic Arabidopsis plants, which exhibited greater root length, chlorophyll content, fresh weight, and survival rate than wild type plants, all in the presence of NaCl. In transgenic plants, hydrogen peroxide (H₂O₂) accumulation was reduced under NaCl stress and loss of chlorophyll content was reduced under H₂O₂ stress. Overall results suggest that dnaK2s from Sy6803 and Sy6906 confer salt and oxidative tolerance in transgenic plants by reduction of H₂O₂ accumulation.     

Somatic embryogenesis and plant regeneration in zygotic embryo explant cultures of rugosa rose

Rugosa rose (Rosa rugosa) is cultivated as a garden flower and an important genetic resource for the breeding of roses (R. hybrida). This study describes culture conditions for high frequency plant regeneration from zygotic embryo explants via somatic embryogenesis in rugosa rose. Mature zygotic embryo, cotyledon, and radicle explants formed embryogenic calluses at frequencies of 38, 6.7, and 8.8% when cultured on half-strength Murashige and Skoog medium (½MS) supplemented with 2.26, 9.05, and 9.05 μM 2,4-dichlorophenoxyacetic acid, respectively. Embryogenic calluses produced numerous somatic embryos, which then developed into plantlets on ½MS without growth regulators. Regenerated plantlets were grown to whole plants in a growth chamber.