Differential response to paraquat induced oxidative stress in two rice cultivars on antioxidants and chlorophyll a fluorescence

The responses of antioxidative system and photosystem II photochemistry of rice (Oryza sativa L.) to paraquat induced oxidative stress were investigated in a chilling-tolerant cultivar Xiangnuo no. 1, and a chilling-susceptible cultivar, IR-50. Electrolyte leakage and malondialdehyde (MDA) content of Xiangnuo no. 1 were little affected by paraquat, but they increased in IR-50. After paraquat treatment, superoxide dismutase (SOD) activity remained high in Xiangnuo no. 1, while it declined in IR-50. Activities of catalase (CAT), ascorbate peroxidase (APX) and glutathione reductase (GR) declined with oxidative stress in both cultivars, but Xiangnuo no. 1 had higher GR activity than IR-50. Under paraquat induced oxidative stress, ascorbic acid (AsA) and reduced glutathione (GSH) concentrations remained high in Xiangnuo no. 1, but decreased in IR-50. The results indicated that higher activities of SOD and GR and higher contents of AsA and GSH in Xiangnuo no. 1 under paraquat induced oxidative stress were associated with its tolerance to paraquat, while paraquat induced damage to IR-50 was related to decreased activities of SOD, APX and GR and contents of AsA and GSH. F _v/F _m, Φ _PSII, and qP remained high in Xiangnuo no. 1, while they decreased greatly in IR-50 under paraquat induced oxidative stress.     

Reductive transformation of parathion and methyl parathion by Bacillus sp.

Based on the results of phenotypic features, phylogenetic similarity of 16S rRNA gene sequences and BIOLOG test, a soil bacterium was identified as Bacillus sp. DM-1. Using either growing cells or a cell-free extract, it transformed parathion and methyl parathion to amino derivatives by reducing the nitro group. Pesticide transformation by a cell-free extract was specifically inhibited by three nitroreductase inhibitors, indicating the presence of nitroreductase activity. The nitroreductase activity was NAD(P)H-dependent, O₂-insensitive, and exhibited the substrate specificity for parathion and methyl parathion. Reductive transformation significantly decreased the toxicity of pesticides.     

Phytase expression in transgenic soybeans: stable transformation with a vector-less construct

A minimal linear gene cassette (35S-phytase gene-nos) with T-DNA borders was acquired by PCR and directly introduced into soybean through the pollen tube pathway. A total of 13% of T₁ plants were positive for phyA by specific PCR. Southern blot analyses showed that phyA insertions were harbored stably in T₂ progeny. Phytase expression level increased 2.5-fold over a 6-week period; its highest activity was 150 U/mg protein, compared to 56 U/mg protein in untransformed controls. Activity of phytase increased to 125 FTU/kg in T₃ transgenic seeds as compared to 64 FTU/kg in wild-type plants.     

Glycosylated trypsin-like proteases from earthworm Eisenia fetida

Although groups of earthworm proteases have been found by several laboratories, it is still unclear how many of the isolated trypsin-like fibrinolytic enzymes are in glycosylated form. Here, eight glycosylated fibrinolytic proteases (EfP-0-1, EfP-0-2, EfP-I-1, EfP-I-2, EfP-II-1, EfP-II-2, EfP-III-1 and EfP-III-2) were isolated from an earthworm species (Eisenia fetida) through a stepwise-purification procedure: ammonium sulfate precipitation, affinity chromatography on a Sepharose-4B column coupled with soybean trypsin inhibitor (SBTI), and ionic chromatography with a DEAE-Cellulose-52 column. Among the eight purified trypsin-like glyco-proteases, EfP-0-2 and EfP-II-2 were newly isolated isozymes. Glycoprotein staining of the proteases on native-PAGE with a Schiff's reagent (sodium meta-periodate) revealed that the eight proteases were glycoproteins. Measurements of the glycan content with sodium meta-periodate and glycoprotein-test reagent showed that these proteases had different carbohydrate contents. Dot-blotting assay with ConA suggested the oligosaccharides were composed of mannose residues.     

Transgene expression in Chinese sweetgum driven by the salt induced expressed promoter

Conventional breeding of Chinese sweetgum is constrained by its long-reproductive cycle, which includes long-juvenile periods, and by its complex reproductive characteristics, including self-incompatibility and a high degree of heterozygosis. Like other tree species, sweetgum has undergone relatively little domestication; the methodology described here in illustrates the possibility of transforming Liquidambar formosana L. obtained from leafy explants using Agrobacter tumefaciens. PCR and Southern blotting show that foreign gene had integrated to genomic DNA. The results indicated that superoxide dismutase (SOD) and peroxidase (POD) activities increased with the stress time in all treated plants and these activities of the transgenic plant were stably higher than those of the control. RT-PCR showed that BADH expressed strongly induced by NaCl. The present study showed that the Rd29A promoter is able to direct osmotant gene expression when plant was exposed to salt, cold, and drought stress, with the advantage that expression was absent or undetectable in natural grow phase.     

Shrinkage of Gobiocypris rarus, Procypris rabaudi, and Sinilabeo rendahli preserved in formalin

Length measurements of preserved fishes are necessary in many types of fish surveys because logistics often do not allow for fish measurement immediately after catch. If the fixative causes significant shrinkage, then the preserved lengths cannot be directly used to indicate accurate live lengths. The objective of this study was to determine how preservation in formalin affects standard length of Gobiocypris rarus larvae (24‐day‐old and newly hatched), larval Procypris rabaudi (4‐day‐old), and larval Sinilabeo rendahli (12‐day‐old). Fishes were measured (to nearest 0.01 mm) and individually fixed in the appropriate formalin solution (2.5% or 5.0% formalin), then re‐measured at 0.5, 1, 3, 7, 14, 30, 45 and 75 days after preservation to follow the time course of shrinkage. Most of the shrinkage occurred within the first half day after preservation. The 5.0% formalin caused a higher relative shrinkage rate than did the 2.5% solution; however, the difference was not statistically significant. In G. rarus, initial shrinkage of newly hatched larvae was higher than that of 24‐day‐old larvae.     

Cloning and characterization of a plastidic glycerol 3-phosphate dehydrogenase cDNA from Dunaliella salina

A cDNA encoding a nicotinamide adenine dinucleotide (NAD+) -dependent glycerol 3-phosphate dehydrogenase (GPDH) has been cloned by rapid amplification of cDNA ends from Dunaliella salina. The cDNA is 3032 base pairs long with an open reading frame encoding a polypeptide of 701 amino acids. The polypeptide shows high homology with published NAD+ -dependent GPDHs and has at its N-terminal a chloroplast targeting sequence. RNA gel blot analysis was performed to study GPDH gene expression under different conditions, and changes of the glycerol content were monitored. The results indicate that the cDNA may encode an osmoregulated isoform primarily involved in glycerol synthesis. The 701-amino-acid polypeptide is about 300 amino acids longer than previously reported plant NAD+ -dependent GPDHs. This 300-amino-acid fragment has a phosphoserine phosphatase domain. We suggest that the phosphoserine phosphatase domain functions as glycerol 3-phosphatase and that, consequently, NAD+ -dependent GPDH from D. salina can catalyze the step from dihydroxyacetone phosphate to glycerol directly. This is unique and a possible explanation for the fast glycerol synthesis found in D. salina.     

A crucial role for dendritic cell (DC) IL-10 in inhibiting successful DC-based immunotherapy: superior antitumor immunity against hepatocellular carcinoma evoked by DC devoid of IL-10

The dendritic cell (DC)-based tumor immunotherapy has been a new promise of cure for cancer patients, but animal studies and clinical trials have thus far only shown limited success, especially in treating established tumors. Certain immunosuppressive mechanisms triggered by tumor cells or the derivatives are believed to be a major obstacle. We studied the role of DC-derived IL-10 and its negative impact on vaccine efficacy in mouse models. Liver tumor cells were injected via the portal vein, giving rise to disseminated intrahepatic tumors, or s.c. to form solid but extrahepatic tumors. Bone marrow-derived DCs were generated from normal or IL-10-deficient mice and used as the vector to deliver tumor Ags. We demonstrate here that DCs devoid of IL-10, a potent immunosuppressive cytokine, are superior over conventional DCs in triggering antitumor immunity. The IL-10(-/-)DCs were highly immunogenic, expressed enhanced levels of surface MHC class II molecules, and secreted increased amounts of Th1-related cytokines. By inducing tumor-specific killing and through the establishment of immunological memory, the vaccines delivered by IL-10(-/-)DCs could evoke strong therapeutic and protective immunity against hepatocellular carcinoma in the mouse models. These findings will have great clinical impact once being translated into the treatment of malignant, and potentially infectious, diseases in humans.     

Phylogeny and biogeography of Cedrus (Pinaceae) inferred from sequences of seven paternal chloroplast and maternal mitochondrial DNA regions

BACKGROUND AND AIMS: Cedrus (true cedars) is a very important horticultural plant group. It has a disjunct distribution in the Mediterranean region and western Himalaya. Its evolution and biogeography are of great interest to botanists. This study aims to investigate the phylogeny and biogeography of Cedrus based on sequence analyses of seven cytoplasmic DNA fragments. METHODS: The methods used were PCR amplification and sequencing of seven paternal cpDNA and maternal mtDNA fragments, parsimony and maximum likelihood analyses of the DNA dataset, and molecular clock estimate of divergence times of Cedrus species. KEY RESULTS: Phylogenies of Cedrus constructed from cpDNA, mtDNA and the combined cp- and mt-DNA dataset are identical in topology. It was found that the Himalayan cedar C. deodara diverged first, and then the North African species C. atlantica separated from the common ancestor of C. libani and C. brevifolia, two species from the eastern Mediterranean area. Molecular clock estimates suggest that the divergence between C. atlantica and the eastern Mediterranean clade at 23.49 +/- 3.55 to 18.81 +/- 1.25 Myr and the split between C. libani and C. brevifolia at 7.83 +/- 2.79 to 6.56 +/- 1.20 Myr. CONCLUSIONS: The results, combined with palaeogeographical and palaeoecological information, indicate that Cedrus could have an origin in the high latitude area of Eurasia, and its present distribution might result from vicariance of southerly migrated populations during climatic oscillations in the Tertiary and further fragmentation and dispersal of these populations. It is very likely that Cedrus migrated into North Africa in the very late Tertiary, while its arrival in the Himalayas would not have been before the Miocene, after which the phased or fast uplift of the Tibetan plateau happened.     

农杆菌介导GUS基因对多年生黑麦草转化的研究

通过检测愈伤组织中GUS基因的瞬间表达,研究农杆菌LBA4404/pCAMBIA1301介导多年生黑麦草的转化体系。通过对多年生黑麦草瞬间表达率的比较,确立了其遗传转化的最佳优化条件。研究发现,多年生黑麦草不同品种的转化率在25%~45%之间变化。多年生黑麦草遗传转化最佳优化条件是预培养10d的胚性愈伤组织、浓度为0.5~0.8OD的农杆菌菌液以及2d共培养时间。在共培养基中添加100μmol/L乙酰丁香酮能有效地提高植物瞬间表达率。两种侵染处理方法比较结果为滤纸滴加法比浸泡法更优。转化后对愈伤组织的干燥处理能抑制农杆菌过度繁殖,能改善愈伤状态,有利于提高转化率。