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Now showing items 1 - 16 of 32

  • Hyperuricemia is associated with impaired intestinal permeability in mice

    Xu, Daxing   Lv, Qiulan   Wang, Xiaofeng   Cui, Xuena   Zhao, Peng   Yang, Xiaomin   Liu, Xiu   Yang, Wan   Yang, Guanpin   Wang, Guangtao   Wang, Pengjun   Wang, Zenglan   Li, Zhiyuan   Xing, Shichao  

    Hyperuricemia is associated with many metabolic diseases. However. the underlying mechanism remains unknown. The gut microbiota has been demonstrated to play significant roles in the immunity and metabolism of the host. In the present study, we constructed a hyperuricemic mouse model to investigate whether the metabolic disorder caused by hyperuricemia is related to intestinal dysbiosis. A significantly increased intestinal permeability was detected in hyperuricemic mice. The difference in microflora between wild-type and hyperuricemic mice accompanies the translocation of gut microbiota to the extraintestinal tissues. Such a process is followed by an increase in innate immune system activation. We observed increased LPS and TNF-alpha levels in the hyperuricemic mice, indicating that hyperuricemic mice were in a state of low-grade systemic inflammation. In addition, hyperuricemic mice presented early injury of parenteral tissue and disordered lipid metabolism. These findings suggest that intestinal dysbiosis due to an impaired intestinal barrier may be the key cause of metabolic disorders in hyperuricemic mice. Our findings should aid in paving a new way of preventing and treating hyperuricemia and its complications. NEW & NOTEWORTHY Hyperuricemia is associated with many metabolic diseases. However, the underlying mechanism remains unknown. We constructed a hyperuricemic mouse model to explore the relationship between intestinal dysbiosis and metabolic disorder caused by hyperuricemia.
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  • Potential biodiesel-producing microalgae:two new strains of Amphikrikos sp.

    Han, Jichang   Zhang, Lin   Wang, Pu   Wang, Song   Yang, Guanpin   Zhao, Lu   Pan, Kehou  

    BH32 and BH43 were isolated and identified as Amphikrikos sp. based on morphological characteristics and phylogenetic trees. They shared the same 18S rRNA sequences, but possessed different amounts of introns in their 18S rDNAs. The specific growth rate, cellular component and fatty acid profile under different growth stages have been surveyed. Their main fatty acids were found to be C16 and C18 groups. Our results indicated that both of them could be taken as excellent candidates of biodiesel. Moreover, the intra-genus polymorphism of 18S rDNA caused by intron presence/absence can provide valuable information for the study of intron evolution.
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  • Co-culturing bacteria and microalgae in organic carbon containing medium.

    Han, Jichang   Zhang, Lin   Wang, Song   Yang, Guanpin   Zhao, Lu   Pan, Kehou  

    BACKGROUND: Microalgae frequently grow in natural environment and long-term laboratory cultures in association with bacteria. Bacteria benefit the oxygen and extracellular substances generated by microalgae, and reimburse microalgae with carbon dioxide, vitamins and so on. Such synergistic relationship has aided in establishing an efficient microalga-bacterium co-culturing mode. Obviously, the mutually beneficial relationship can be strengthened with the increase of the densities of microalgae and bacteria. However, nearly all of the early co-cultures were performed under photoautotrophic conditions, thus both microalgae and bacteria were at relatively low densities. In this study, the feasibility of bacteria-microalgae co-cultured under mixotrophic conditions was studied.; RESULTS: Firstly, bacteria mingled with xenic microalgae were isolated and identified based on their 16S rRNA gene sequence (16S rDNA hereafter). Then, the two most frequently found strains of Muricauda sp. were co-cultured with axenic microalga (Tetraselmis chuii, Cylindrotheca fusiformis and Nannochloropsis gaditana) in extra organic carbon containing medium. At the end of a co-culture period of 33days, we found that the final cell density of T. chuii and C. fusiformis of various treatments was remarkably higher than that of controls (21.37-31.18 and 65.42-83.47%, respectively); on the contrary, the growth of N. gaditana was markedly inhibited. During the co-culture of bacteria with C. fusiformis, the cell density of two strains of bacteria firstly decreased, then increased and maintained at a relatively steady level. However, the cell density of bacteria performed a sustaining downward trend when they were co-cultured with T. chuii and N. gaditana.; CONCLUSIONS: Our findings proved that microalgae-bacteria co-cultures under mixotrophic conditions are quite effective strategy for microalgal cultivation.=20
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  • Identification of Microplankton in South China Sea with Image-Matching Individual PCR

    Li, Si   Yang, Guanpin   Zhu, Baohua   Pan, Kehou  

    In this study, marine microplankton were identified by combining standard light microscopy with Sanger 18S rRNA gene sequencing. The image-matching individual PCR technique was applied to identify the image collectable unicellular microplankton to genera. Instead of pure strain culture and morphological identification, microplankton individual cells were isolated and fixed with glutaraldehyde, frozen and stored for months. Finally, they were imaged under a microscope and molecularly identified via phylogenetic analysis of their 18S ribosomal RNA gene (18S rDNA). Microplankton cells were collected at 30 locations in South China Sea, and were assigned to 21 known and 4 unidentified genera (2 uncultured fungi and 2 uncultured stramenopiles) with phylogenetic analysis in parallel to the morphological identification.
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  • Identification of Microplankton in South China Sea with Image-Matching Individual PCR

    Li, Si   Yang, Guanpin   Zhu, Baohua   Pan, Kehou  

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  • Mutational analysis of the zinc metalloprotease EmpA of Vibrio anguillarum.

    Yang, Hui   Chen, Jixiang   Yang, Guanpin   Zhang, Xiao-Hua   Li, Yun  

    The extracellular zinc metalloprotease, EmpA, is a putative virulence factor involved in pathogenicity of the fish pathogen Vibrio anguillarum. The 611-amino acid precursor of this enzyme is encoded by the empA gene. The residues His346, His350, Glu370, Glu347, His429, Tyr361 and Asp417 are highly conserved and putatively function together at the active site of the enzyme. In this study, empA was inserted into pET24d(+) and expressed in Escherichia coli strain BL21(DE3) as a 6 x His tagged protein (r-EmpA). All the conserved residues of EmpA mentioned above were individually mutated by site-directed mutagenesis and the mutants were also expressed (m-r-EmpAs). r-EmpA and m-r-EmpAs were purified, and assayed for their proteolytic activities with azocasein as the substrate and cytotoxicities on a flounder gill cell line. m-r-EmpAs that had been mutated at His346, His350, Glu370 and Glu347 almost completely lost their proteolytic activity and cytotoxicity, pointing towards the essential roles played by these residues. In contrast, those mutated at Tyr361, His429 and Asp417 still retained a partial proteolytic activity and cytotoxicity. Our results indicate that these conserved residues play important roles in enzymatic activity and that the proteolytic activity of the enzyme is involved in the pathogenesis of V. anguillarum
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  • Genetic Transformation of Tribonema minus,a Eukaryotic Filamentous Oleaginous Yellow-Green Alga

    Zhang, Yan   Wang, Hui   Yang, Ruigang   Wang, Lihao   Yang, Guanpin   Liu, Tianzhong  

    Eukaryotic filamentous yellow-green algae from the Tribonema genus are considered to be excellent candidates for biofuels and value-added products, owing to their ability to grow under autotrophic, mixotrophic, and heterotrophic conditions and synthesize large amounts of fatty acids, especially unsaturated fatty acids. To elucidate the molecular mechanism of fatty acids and/or establish the organism as a model strain, the development of genetic methods is important. Towards this goal, here, we constructed a genetic transformation method to introduce exogenous genes for the first time into the eukaryotic filamentous alga Tribonema minus via particle bombardment. In this study, we constructed pSimple-tub-eGFP and pEASY-tub-npt? plasmids in which the green fluorescence protein (eGFP) gene and the neomycin phosphotransferase ?-encoding G418-resistant gene (npt?) were flanked by the T. minus-derived tubulin gene (tub) promoter and terminator, respectively. The two plasmids were introduced into T. minus cells through particle-gun bombardment under various test conditions. By combining agar and liquid selecting methods to exclude the pseudotransformants under long-term antibiotic treatment, plasmids pSimple-tub-eGFP and pEASY-tub- npt? were successfully transformed into the genome of T. minus, which was verified using green fluorescence detection and the polymerase chain reaction, respectively. These results suggest new possibilities for efficient genetic engineering of T. minus for future genetic improvement.
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  • Identification of barley varieties used in beer production by microsatellite DNA markers

    Lin, Yan   Liao, Meijie   Yang, Guanpin   Yu, Wengong   Guan, Huashi   Fan, Wei   Dong, Jianjun  

    Microsatellite DNA (or simple sequence repeat [SSR]) markers were used in this study for the identification of barley varieties. Seventeen SSRs were selected from a representative, highly informative genotyping set of barley SSRs. These SSRs detected polymorphism among 17 barley varieties imported by Tsingtao Brewery Co. Ltd. and used in its beer production. The number of alleles at different loci varied from two to five. Of 17 varieties, seven could be distinguished from each other and from the remaining varieties using specific alleles. As an example, the loci Bmag0125, Bmag0120, HVM62, and Bmag0353 in combination were able to distinguish all barley varieties tested. An allele-based retrieval system was developed for the 17 barley varieties based on these four SSRs.
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  • The immune responses in Chinese mitten crab Eriocheir sinensis challenged with double-stranded RNA

    Dong, Chaohua   Zhao, Jianmin   Song, Linsheng   Wang, Lingling   Qiu, Limei   Zheng, Peilin   Li, Ling   Gai, Yuncao   Yang, Guanpin  

    Double-stranded RNA (dsRNA) is a virus-associated molecular pattern which induces antiviral innate immune responses and RNA interference (RNAi) in mammals. In invertebrates, RNAi phenomenon has been widely studied, but dsRNA-induced innate immune response is seldom reported. In the present study, two different dsRNAs specific for green fluorescent protein (GFP) and the putative D1 protein of photosystem II (NoPSD) from Nannochloropsis oculata, were employed to challenge Chinese mitten crab Eriocheir sinensis. The temporal changes of phenoloxidase (PO), acid phosphatase (ACP), superoxide dismutase (SOD) and malondialdehyde (MDA) content, as well as the mRNA expression of some immune-related genes were examined in order to estimate the effect of dsRNAs on the innate immunity of E. sinensis. The activities of PO, ACP and SOD significantly increased after dsRNA treatment, whereas malondialdehyde (MDA) content did not change significantly. Among the examined genes, only the mRNA expression of EsALF, an antibacterial peptide in E. sinensis, was significantly up-regulated (about 5 fold, P < 0.05) at 12 h after dsRNA treatment, while no significant expression changes were observed among the other immune genes. The increase of PO, ACP and SOD activities, and mRNA expression level of EsALF after dsRNA stimulation indicate that phenoloxidase, hydrolytic enzyme, antioxidation and EsALF were involved in dsRNA-induced innate immunity, suggesting that broad-spectrum immune responses could be induced by dsRNA in E. sinensis. (C) 2009 Elsevier Ltd. All rights reserved.
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  • The involvement of HSP22 from bay scallop Argopecten irradians in response to heavy metal stress

    Zhang, Lei   Zhao, Jianmin   Qiu, Limei   Dong, Chaohua   Li, Fengmei   Zhang, Huan   Yang, Guanpin  

    Heat shock protein 22 (HSP22) is an important member of small heat shock protein (sHSP) subfamily which plays a key role in the process of protecting cells, facilitating the folding of nascent peptides, and responding to stress. In the present study, the cDNA of HSP22 was cloned from Argopecten irradians (designated as AiHSP22) by rapid amplification cDNA end (RACE) based on the expressed sequence tags (ESTs). The full-length cDNA of AiHSP22 was of 1,112 bp, with an open reading frame of 588 bp encoding a polypeptide of 195 amino acids. The deduced amino acid sequence of AiHSP22 showed high similarity to previously identified HSP22s. The expression patterns of AiHSP22 mRNA in different tissues and in haemocytes of scallops exposed to Cd2+, Pb2+ or Cu2+ were investigated by real-time quantitative RT-PCR. The mRNA of AiHSP22 was constitutively expressed in all examined tissues, including haemocyte, muscle, kidney, gonad, gill and heart. The expression level in heart and muscle was higher than that in other tissues. The mRNA level of AiHSP22 in haemocytes was up-regulated after a 10 days exposure of scallops to Cu2+, Pb2+ and Cd2+. However, the expression of AiHSP22 did not increase linearly along with the rise of heavy metal concentration. Different concentrations of the same metal resulted in different effects on AiHSP22 expression. The sensitive response of AiHSP22 to Cu2+, Pb2+ and Cd2+ stress indicated that it could be developed as an indicator of exposure to heavy metals for the pollution monitoring programs in aquatic environment.
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  • Analysis of expressed sequence tags from the marine microalga Nannochloropsis oculata (Eustigmatophyceae)

    Shi, Juan   Yu, Jianzhong   Zhu, Baohua   Yang, Guanpin   Yu, Wengong   Zhang, Xinyu  

    Nannochloropsis oculata (Droop) D. J. Hibberd (Eustigmatophyceae), a marine eukaryotic unicellular alga, is widely used in mariculture as live feed. It is considered to be of high nutritional value owing to its high content of proteins; polyunsaturated fatty acids (PUFAs), especially eicosapentaenoic acid (EPA, C20:5n3); and diverse pigments. Previous studies of this microalga focused on its taxonomy, culture, and biochemistry, but little is known at the molecular level. Establishing a molecular base is vital to understand the biological processes of this alga. Therefore, we constructed a cDNA library using algal cells grown at exponential growth phase and carried out expressed sequence tag (EST) analysis. A total of 1,960 nonredundant sequences (NRSs) were generated for N. oculata clone CS-179. Only 32.5% of NRSs showed significant similarity (E < 1e-04) to proteins registered in the GenBank nonredundant protein database. The KOG (clusters of euKaryotic Orthologous Groups) profile database returned significant hits for 490 NRSs. Analysis revealed that a large proportion of NRSs could be unique to this microalga.
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  • Development of 15 polymorphic genic microsatellite DNA markers of Pacific abalone Haliotis discus hannai

    Sun, Xiuqing   Zhen, Minggang   Yang, Guanpin  

    Fifteen polymorphic genic microsatellite DNA markers were developed based on the expressed sequence tags of Pacific abalone Haliotis discus hannai we ourselves generated, which were used to type 32 individuals representing a cultured population. The number of alleles each locus ranged from two to 12. The observed and expected heterozygosities ranged from 0.094 to 0.969 and from 0.091 to 0.878, respectively. Among 15 loci, four were found to deviate significantly from Hardy-Weinberg equilibrium (P-HW < 0.001). No linkage disequilibrium was found between these loci. It is certain that these markers will facilitate the management and exploitation of the genetic resource of Pacific abalone.
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  • Construction of genetic linkage maps of guppy (Poecilia reticulata) based on AFLP and microsatellite DNA markers

    Shen, Xueyan   Yang, Guanpin   Liu, Yongjian   Liao, Meijie   Wang, Xiaochen   Zhu, Mingzhuang   Song, Weibo   Zou, Guiwei   Wei, Qiwei   Wang, Dengqiang   Chen, Daqing  

    Genetic linkage maps of guppy (Poecilia reticulata) were constructed using microsatellite DNA and AFLP markers and a pseudo-testcross mapping strategy. (AC)(n) microsatellite DNA markers were developed and used to genotype sixty-one full-sib progenies (F-1) and their two parents. Of 293 microsatellite DNA markers, 101 segregated at 1:1 or 1:1:1:1 ratios. In addition, 336 AFLP markers segregated also in F-1 progenies at 1:1 ratio, which were produced using 91 primer combinations. All these markers were mapped with two linkage maps produced, one each parent. Female map included 135 markers in 22 linkage groups, covering a total of 1267.7 cM in length. Male map included 172 markers in 20 linkage groups, covering a total of 1771.2 cM in length. (C) 2007 Elsevier B.V. All rights reserved.
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  • Isolation and characterization of 22 polymorphic microsatellite DNA markers of Japanese sea bass (Laterolabrax japonicus)

    Jiang, Xin   Liao, Meijie   Liu, Yongjian   Gao, Tianxiang   Yang, Guanpin  

    Japanese sea bass (Laterolabrax japonicus) inhabits the coast of East Asia and is cage-cultured currently in China as well. Twenty-two microsatellite DNA markers were developed and used to type 35 individuals collected along the Chinese coast. The number of alleles per locus ranged from three to 23. The expected heterozygosity ranged from 0.111 to 0.938, while the observed heterozygosity ranged from 0.114 to 0.914. All 22 loci are neutral and independent of each other; two deviated significantly from Hardy-Weinberg equilibrium. These microsatellite DNA markers are moderately informative, which will certainly facilitate the management and exploitation of the genetic resource of L. japonicus.
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  • Characterization and pathogenicity of the zinc metalloprotease empA of Vibrio anguillarum expressed in Escherichia coli.

    Yang, Hui   Chen, Jixiang   Yang, Guanpin   Zhang, Xiao-Hua   Li, Yun   Wang, Min  

    The extracellular zinc metalloprotease (EmpA) is a putative pathogenic factor involved in the invasive process of the fish pathogen Vibrio anguillarum. It is synthesized as a 611-amino-acid preproprotease. The gene encoding EmpA (empA) has already been cloned and sequenced. In this study, empA was inserted into pET24d(+) and expressed in Escherichia coli BL21(DE3). Recombinant EmpA with His-tag was purified in a single step with a His-binding Ni-affinity column to a purity >95%. In addition, proteolytic activity, cytotoxicity, fish pathogenicity, and solubility of the recombinant protein were determined.
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  • Involvement of peroxinectin in the defence of red swamp crayfish Procambarus clarkii against pathogenic Aeromonas hydrophila

    Dong, Chaohua   Wei, Zhu   Yang, Guanpin  

    Cell adhesion factors are important immune components for invertebrate to immobilize, phagocytose or encapsulate invasive microorganisms and foreign particles. In this study, a new cell adhesion factor, peroxinectin (refered as Pcpxin) was isolated from hemocytes of red swamp crayfish (Procambarus clarkii). The full-length cDNA of Pcpxin was 3014 bp encoding a protein of 819 amino acid residues with a predicted molecular weight of 89.0 kDa and a calculational isoelectric point of 6.93. The putative amino acid sequence contained a peroxidase domain and a signal peptide of 21 amino acid residues, and exhibited high identity to peroxinectin from Pacifastacus leniusculus (85%), Fenneropenaeus chinensis (62%) and Scylla serrata (58%), as well as peroxidase from Camponotus floridanus (40%), Pediculus humanus corporis (39%), and Culex quinquefasciatus (38%). Quantitative real time PCR revealed that mRNA expression of Pcpxin in hemocytes could be inhibited by challenge with heat-killed Aeromonas hydrophila, suggesting that Pcpxin was involved in immune responses to A. hydrophila. RNA interference (RNAi) experiment demonstrated that silencing Pcpxin significantly reduced the survival rate of red swamp crayfishes after challenge with A. hydrophila, which indicated that Pcpxin was important for P. clarkii to survive A. hydrophila infection. Moreover, silencing Pcpxin inhibited the up-regulation of crustin1 and lysozyme expression in response to challenge with heat-killed A. hydrophila. This result suggested that Pcpxin might participate in antibacterial peptide gene expression and thereby might be involved in signal transduction pathway regulating the expression of antibacterial peptide gene. (C) 2011 Elsevier Ltd. All rights reserved.
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