Multiplex polymerase chain reaction/membrane hybridization assay for detection of genetically modified organisms

To improve detection efficiency and result accuracy, four screening primer pairs, four identifying primer pairs, one common primer pair and corresponding probes were designed for the development of multiplex polymerase chain reaction/membrane hybridization assay (MPCR-MHA) for detection of the foreign genes insert in genetically modified organisms (GMOs). After detecting condition and parameter were optimized and determined, MPCR reactions were developed for amplifying several target genes simultaneously in one tube. Primers were labeled with biotin at the 5'-end; biotinylated MPCR products were detected by hybridization to the oligonucleotide probes immobilized on a membrane with subsequent colorimetric detection to confirm hybridization. The testing of screening primers can judge whether the sample contains GMOs, and that of identifying primers can further judge what kinds of trait genes are contained in the sample. We detected nine soybean samples, six maize samples, seven potato samples and two rice samples by the MPCR-MHA method; at the same time we also detected them with single PCR-MHA method. The results between two methods have good consistency.     

The PPAR-alpha activator fenofibrate fails to provide myocardial protection in ischemia and reperfusion in pigs

Rodent studies suggest that peroxisome proliferator-activated receptor-alpha (PPAR-alpha) activation reduces myocardial ischemia-reperfusion (I/R) injury and infarct size; however, effects of PPAR-alpha activation in large animal models of myocardial I/R are unknown. We determined whether chronic treatment with the PPAR-alpha activator fenofibrate affects myocardial I/R injury in pigs. Domestic farm pigs were assigned to treatment with fenofibrate 50 mg.kg(-1).day(-1) orally or no drug treatment, and either a low-fat (4% by weight) or a high-fat (20% by weight) diet. After 4 wk, 66 pigs underwent 90 min low-flow regional myocardial ischemia and 120 min reperfusion under anesthetized open-chest conditions, resulting in myocardial stunning. The high-fat group received an infusion of triglyceride emulsion and heparin during this terminal experiment to maintain elevated arterial free fatty acid (FFA) levels. An additional 21 pigs underwent 60 min no-flow ischemia and 180 min reperfusion, resulting in myocardial infarction. Plasma concentration of fenofibric acid was similar to the EC50 for activation of PPAR-alpha in vitro and to maximal concentrations achieved in clinical use. Myocardial expression of PPAR-alpha mRNA was prominent but unaffected by fenofibrate treatment. Fenofibrate increased expression of carnitine palmitoyltransferase (CPT)-I mRNA in liver and decreased arterial FFA and lactate concentrations (each P < 0.01). However, fenofibrate did not affect myocardial CPT-I expression, substrate uptake, lipid accumulation, or contractile function during low-flow I/R in either the low- or high-fat group, nor did it affect myocardial infarct size. Despite expression of PPAR-alpha in porcine myocardium and effects of fenofibrate on systemic metabolism, treatment with this PPAR-alpha activator does not alter myocardial metabolic or contractile responses to I/R in pigs.     

The miR156‐SPL9‐DFR pathway coordinates the relationship between development and abiotic stress tolerance in plants

Young organisms have relatively strong resistance to diseases and adverse conditions. When confronted with adversity, the process of development is delayed in plants. This phenomenon is thought to result from the rebalancing of energy, which helps plants to coordinate the relationship between development and stress tolerance; however, the molecular mechanism underlying this phenomenon remains mysterious. In this study, we found that miR156 integrates environmental signals to ensure timely flowering, thus enabling the completion of breeding. Under stress conditions, miR156 is induced to maintain the plant in the juvenile state for a relatively long period of time, whereas under favorable conditions, miR156 is suppressed to accelerate the developmental transition. Blocking the miR156 signaling pathway in Arabidopsis thaliana with 35S::MIM156 (via target mimicry) increased the sensitivity of the plant to stress treatment, whereas overexpression of miR156 increased stress tolerance. In fact, this mechanism is also conserved in Oryza sativa (rice). We also identified downstream genes of miR156, i.e. SQUAMOSA PROMOTER BINDING PROTEIN‐LIKE 9 (SPL9) and DIHYDROFLAVONOL‐4‐REDUCTASE (DFR), which take part in this process by influencing the metabolism of anthocyanin. Our results uncover a molecular mechanism for plant adaptation to the environment through the miR156‐SPLs‐DFR pathway, which coordinates development and abiotic stress tolerance.     

Insecticidal activity and chemical composition of the essential oil of Artemisia vestita from China against Sitophilus zeamais

In our screening program for new agrochemicals from local wild plants, essential oil of Artemisia vestita Wall (Asteraceae) was found to possess strong insecticidal activity against maize weevil, Sitophilus zeamais Motsch. Essential oil of aerial parts of A. vestita was obtained from hydrodistillation and was investigated by GC and GC–MS. The main components of essential oil were grandisol (40.29%), 1,8-cineol (14.88%) and camphor (11.37%). The essential oil of A. vestita possessed strong fumigant toxicity against S. zeamais adults with a LC₅₀ value of 13.42 mg/L air. The essential oil of A. vestita also showed contact toxicity against S. zeamais adults with a LD₅₀ value of 50.62 mg/adult.     

Phytoextraction of heavy metals from contaminated soil by co-cropping with chelator application and assessment of associated leaching risk

Phytoextraction using hyperaccumulating plants is generally time-consuming and requires the cessation of agriculture. We coupled chelators and a co-cropping system to enhance phytoextraction rates, while allowing for agricultural production. An experiment on I m3 lysimeter beds was conducted with a co-cropping system consisting of the hyperaccumulator Sedum alfredii and low-accumulating corn (Zea Mays, cv. Huidan-4), with addition ofa mixture of chelators (MC), to assess the efficiency of chelator enhanced co-crop phytoextraction and the leaching risk caused by the chelator. The results showed that the addition of MC promoted the growth of S. alfredii in the first crop (spring-summer season) and significantly increased the metal phytoextraction. The DTPA-extractable and total metal concentrations in the topsoil were also reduced more significantly with the addition of MC compared with the control treatments. However, mono-cropped S. alfredii without MC was more suitable for maximizing S. alfredii growth and therefore phytoextraction of Zn and Cd during the autumn-winter seasons. No adverse impact to groundwater due to MC application was observed during the experiments with three crops and three MC applications. But elevated total Cd and Pb concentrations among subsoils compared to the initial subsoil concentrations were found for the co-crop + MC treatment after the third crop.     

Effects of phorbol esters on insulin receptor function and insulin action in hepatocytes: evidence for heterogeneity

We investigated the effect of phorbol 12-myristate 13-acetate (PMA), a protein kinase C (PKC) activator on insulin receptors and insulin action in freshly isolated and primary cultures of rat hepatocytes. PMA (1 x 10^–7 M) did not alter insulin receptor numbers or affinity either acutely or chronically but within 60 minute inactivated insulin stimulated tyrosine kinase of the insulin receptor. PKC activation inhibitied insulin (1 x 10^–7M) stimulation of glycogen and lipid synthesis with a decrease or no change in basal glycogenesis and lipogenesis respectively. However, PKC activation did not alter insulin stimulated or basal amino acid transport even though PCK activation inhibited insulin stimulation of the insulin. receptor tyrosine kinase. Thus, within one tissue, PKC activation has differential effect on insulin action depending on which pathway is examined. Furthermore, insulin stimulation of the insulin receptor tyrosine kinase may not be a necessary step for all insulin signaling pathways.     

Aureonitols A and B,Two New C13‐Polyketides from Chaetomium globosum,an Endophytic Fungus in Salvia miltiorrhiza

Two new C₁₃‐polyketides, aureonitols A and B ( 1 and 2 ), along with five known compounds ( 3 – 7 ), were isolated from the solid fermentation culture of the plant endophytic fungus Chaetomium globosum from the aerial parts of Salvia miltiorrhiza. The structures and absolute configurations of 1 and 2 were determined by comprehensive spectroscopic data analysis and computed methods. Compound 5 was found to display the remarkable antimicrobial activities against four multidrug‐resistant bacteria (Enterococcus faecalis, Enterococcus faecium, Staphylococcus aureus, and Staphylococcus epidermidis) with MIC values of 3.13–6.25 μg/mL (ciprofloxacin: 0.78–1.56 μg/mL), and also against all tested fungal strains with MIC values of 3.13–25 μg/mL (ketoconazole: 0.78–12.50 μg/mL).     

Human serum and glucocorticoid-inducible kinase-like kinase (SGKL) phosphorylates glycogen syntheses kinase 3 beta (GSK-3beta) at serine-9 through direct interaction

Serum and glucocorticoid-inducible kinase-like kinase (SGKL) has been identified as a new integrator that decodes lipid signals produced by the activation of phosphoinositide 3-kinase (PI3K). SGKL is activated via its lipid-binding domain (phox homology domain) in response to PI3K signaling. However, downstream targets of SGKL as well as the role of SGKL as a mediator in PI3K signaling in human tissues remain to be established. In this study, we identified human glycogen synthase kinase 3 beta (GSK-3beta) as a specific interacting partner with SGKL in a yeast two-hybrid screening of human brain cDNA library. The association between these two proteins is confirmed independently in human embryonic kidney (HEK293) cells by co-immunoprecipitation. Furthermore, the kinase activity of wild-type SGKL was required for the in vitro phosphorylation of a GSK-3 crosstide fusion protein at serine-21/9 as demonstrated with a Phospho-GSK-3alpha/beta (Ser21/9) specific antibody. The present results provide strong evidences that SGKL could utilize GSK-3beta as a direct downstream target by phosphorylating GSK-3beta at serine-9.     

Crystal structures of interleukin-2 tyrosine kinase and their implications for the design of selective inhibitors

Interleukin-2 tyrosine kinase, Itk, is an important member of the Tec family of non-receptor tyrosine kinases that play a central role in signaling through antigen receptors such as the T-cell receptor, B-cell receptor, and Fcepsilon. Selective inhibition of Itk may be an important way of modulating many diseases involving heightened or inappropriate activation of the immune system. In addition to an unliganded nonphophorylated Itk catalytic kinase domain, we determined the crystal structures of the phosphorylated and nonphosphorylated kinase domain bound to staurosporine, a potent broad-spectrum kinase inhibitor. These structures are useful for the design of novel, highly potent and selective Itk inhibitors and provide insight into the influence of inhibitor binding and phosphorylation on the conformation of Itk.