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Owing to the high nutritional value and extensive medicinal use of its products, Chinese jujube (Ziziphus jujuba Mill) is one of the most important fruit crops in China. However, jujube fruits are highly perishable and thus have a short shelf life, which is a serious hindrance to the industry. Better understanding of the molecular mechanisms underlying jujube fruit softening is fundamental to overcome the problem. Thus, both forward and reverse suppression subtractive hybridization (SSH) cDNA libraries were constructed to identify differentially expressed genes for fruit at half-red ripening stage and complete red stage. As a result of dot blot confirmation, a total of 154 differentially expressed genes were identified. After removed low-quality regions and screened for vector contamination, blasted with the non-redundant NCBI databases, 78.6 % of sequences exhibited high homology to previously identified or putative proteins. All the ESTs were annotated and classified according to the terms of the three main Gene Ontology vocabularies using the Blast2GO software. Furthermore, the quantitative real-time PCR was carried out for 17 genes to validate the genes differentially expressed from the SSH libraries. And the full-length sequences of galactose oxidase and aldehyde dehydrogenase genes were obtained. It is the first step to explore the functional genomics and regulatory networks during the storage period of jujube fruit. The identification of the genes differentially expressed is helpful to understand the ripening and softening of the jujube fruit at the molecular level.
RAPD (randomly amplified polymorphic DNA) and ISSR (intersimple sequence repeat) fingerprinting on HpaII/MspI-digested genomic DNA of nine elite japonica rice cultivars implies inter-cultivar DNA methylation polymorphism. Using both DNA fragments isolated from RAPD or ISSR gels and selected low-copy sequences as probes, methylation-sensitive Southern blot analysis confirms the existence of extensive DNA methylation polymorphism in both genes and DNA repeats among the rice cultivars. The cultivar-specific methylation patterns are stably maintained, and can be used as reliable molecular markers. Transcriptional analysis of four selected sequences (RdRP, AC9, HSP90 and MMR) on leaves and roots from normal and 5-azacytidine-treated seedlings of three representative cultivars shows an association between the transcriptional activity of one of the genes, the mismatch repair (MMR) gene, and its CG methylation patterns.
Mitochondrial uncoupling proteins play important roles in generation of metabolic thermogenesis, response to stress situations, and regulation of energy metabolism. We demonstrated here that the absence of LeUCP in tomato inhibited photosynthesis using virusinduced gene-silencing approach. A significant decrease in the rate of CO2 assimilation in LeUCP-silencing plants was observed over a range of different light intensities. Absence of LeUCP resulted in lower net photosynthetic rate, lightsaturated rate of the CO2 assimilation (Asat), maximum carboxylation rates (Vcmax) and maximum RuBP regeneration rate (Jmax). Activities of ribulose-1,5-bisphosphate carboxylase/oxygenase Rubisco and stromal fructose-1,6- bisphosphatase and genes expression levels encoded Calvin cycle enzymes of LeUCP gene-silencing plants were inhibited. However, silencing of LeUCP gene had no effect on Fv/Fm, but decreased photochemical quenching and electron transport rate. Meanwhile, non-photochemical quenching and Je (PSII), the distribution of Je (PCR) and Je (PCO), the content of AsA, NAD, and the ratios of NAD?/ NADH, AsA/DHA were significantly reduced with increased reactive oxygen species while GSH and GSSG were unaltered. Taken together, these results strongly suggest that LeUCP gene in tomato leaves is crucial in maintaining the redox poise of the mitochondrial electron transport chain to facilitate photosynthetic metabolism.
Phoebe bournei commonly called nanmu is an important and endemic wood species in China, and its planting, nursing, and preserving are often affected by drought stress. Two-year-old P. bournei seedlings were subjected to water stress and recovery treatment to study their physiological and biochemical responses. Physiological and biochemical indices did not change when seedlings were subjected to mild water stress (˂15 days of water withholding). As drought stress intensified (>20 days of water withholding), malondialdehyde and electrolyte leakage increased, and chlorophyll and soluble protein decreased, indicating an increased oxidative stress induced by water deficit. Enhanced activities of superoxide dismutase (SOD) and peroxidase (POX), accumulation of free proline and total soluble sugar contribute to plant protection against the oxidative stress. However, SOD and POX decreased when seedlings were subjected to an extended drought. After 5 days of recovery, physiological and biochemical indices were not restored to the control level values except for leaf relative water content when the seedlings were subjected to more than 20 days water stress. These results demonstrate that P. bournei could enhance their ability to mitigate water stress effects by up-regulating antioxidant system and osmotic adjustment, but these two protective mechanisms were limited when seedlings were subjected to moderate and severe water stress. The threshold of water deficit to P. bournei seedlings is 15–20 days, and permanent damage will be induced if water status is not improved before this threshold. The results will provide some theoretical and practical guidance for nanmu afforestation and production.
African swine fever is an acute, febrile and highly virulent porcine disease causing serious economic losses worldwide. The pK205R protein of the African swine fever virus (ASFV) is largely expressed in the early stages of infection, which has given the K205R gene extensive attention. In this study, the ASFV K205R was cloned and expressed in Escherichia coli BL21 (DE3). Expression of histidine-tagged pK205R with a molecular mass of 44 kDa was determined by 12% sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blot analysis. Optimisation of culture conditions allowed induction of the recombinant protein with 0.4 mM Isopropyl β-D-thiogalactoside (IPTG) at 37oC for 2 h. The protein existed in cellular supernatant and was purified using a Ni-NTA resin column. The purified protein was used to immunize rabbits four times to enable the production of polyclonal antibodies, and the antiserum titre was detected by ELISA. The results showed that the purified pK205R can react with ASFV positive serum specifically by Western blotting. The pK205R had high antigenicity, which indicated that pK205R could be used as an antigen for detection of ASFV-specific antibodies in ELISA testing, and the recombinant protein could contribute to further research of the action and structure of pK205R.
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