Total chemical synthesis,assembly of human torque teno virus genome

Torque teno virus(TTV)is a nonenveloped virus containing a single-stranded,circular DNA genome of approximately 3.8kb.We completely synthesized the 3808 nucleotides of the TTV(SANBAN isolate)genome,which contains a hairpin structure and a GC-rich region.More than 100 overlapping oligonucleotides were chemically synthesized and assembled by polymerise chain assembly reaction(PCA),and the synthesis was completed with splicing by overlap extension(SOEing).This study establishes the methodological basis of the chemical synthesis of a viral genome for use as a live attenuated vaccine or gene therapy vector.     

Inhibition of mitochondrial complex I improves glucose metabolism independently of AMPK activation

Accumulating evidences showed metformin and berberine, well‐known glucose‐lowering agents, were able to inhibit mitochondrial electron transport chain at complex I. In this study, we aimed to explore the antihyperglycaemic effect of complex I inhibition. Rotenone, amobarbital and gene silence of NDUFA13 were used to inhibit complex I. Intraperitoneal glucose tolerance test and insulin tolerance test were performed in db/db mice. Lactate release and glucose consumption were measured to investigate glucose metabolism in HepG2 hepatocytes and C2C12 myotubes. Glucose output was measured in primary hepatocytes. Compound C and adenoviruses expressing dominant negative AMP‐activated protein kinase (AMPK) α1/2 were exploited to inactivate AMPK pathway. Cellular NAD⁺/NADH ratio was assayed to evaluate energy transforming and redox state. Rotenone ameliorated hyperglycaemia and insulin resistance in db/db mice. It induced glucose consumption and glycolysis and reduced hepatic glucose output. Rotenone also activated AMPK. Furthermore, it remained effective with AMPK inactivation. The enhanced glycolysis and repressed gluconeogenesis correlated with a reduction in cellular NAD⁺/NADH ratio, which resulted from complex I suppression. Amobarbital, another representative complex I inhibitor, stimulated glucose consumption and decreased hepatic glucose output in vitro, too. Similar changes were observed while expression of NDUFA13, a subunit of complex I, was knocked down with gene silencing. These findings reveal mitochondrial complex I emerges as a key drug target for diabetes treatment. Inhibition of complex I improves glucose homoeostasis via non‐AMPK pathway, which may relate to the suppression of the cellular NAD⁺/NADH ratio.     

鸡减蛋综合征病毒（EDSV—76）末端前体蛋白的基因结构分析

从中国发病鸡群中分离的鸡减蛋综合征病毒弱毒株ＡＡ－２,经常规方法提取其病毒核酸后,组建了完整的限制性内切酶ＰｓｔＩ及ＨｉｎｇⅢ水解片段的基因文库,并对其中ＨｉｎｄⅢ,－ＳａｃⅠ进行了序列测定。同源比较分析证明：其Ｌ链含编码病毒末端前体蛋白,容量为５８０个氨基酸残基的开放读码框架。     

Complete genome sequence of the bacterium Ketogulonicigenium vulgare Y25

Ketogulonicigenium vulgare is characterized by the efficient production of 2KGA from L-sorbose. Ketogulonicigenium vulgare Y25 is known as a 2-keto-L-gulonic acid-producing strain in the vitamin C industry. Here we report the finished, annotated genome sequence of Ketogulonicigenium vulgare Y25.     

A novel and highly efficient production system for recombinant adeno-associated virus vector

Recombinant adeno-associated virus(rAAV) has proven to be a promising gene delivery vector for human gene therapy. However, its application has been limited by difficulty in obtaining enough quantities of high-titer vector stocks. In this paper, a novel and highly efficient production system for rAAV is described. A recombinant herpes simplex virus type 1(rHSV-1) designated HSV1-rc/△UL2, which expressed adeno-associated virus type2(AAV-2) Rep and Cap proteins, was constructed previously. The data confirmed that its functions were to support rAAV replication and packaging, and the generated rAAV was infectious. Meanwhile, an rAAV proviral cell line designated BHK/SG2, which carried the green fluorescent protein(GFP) gene expression cassette, was established by transfecting BHK-21 cells with rAAV vector plasmid pSNAV-2-GFP. Infecting BHK/SG2 with HSV1-rc/△UL2 at an MOI of 0.1 resulted in the optimal yields of rAAV, reaching 250 transducing unit(TU) or 4.28×104 particles per cell. Therefore, compared with the conventional transfection method, the yield of rAAV using this "one proviral cell line, one helper virus" strategy was increased by two orders of magnitude. Large-scale production of rAAV can be easily achieved using this strategy and might meet the demands for clinical trials of rAAV-mediated gene therapy.     

中国大鲵研究进展

中国大鲵是我国特有濒危的两栖物种,是研究生物进化、生物多样性、性别决定分子机制等问题的好材料。近年来,人们对它的研究力度不断加大,著述颇多。本文对大鲵的生态保护、解剖发育、生理生化、遗传进化等方面的近期研究资料进行了整理和回顾,也简要探讨了今后大鲵研究的主要工作,以期为研究者们提供有价值的资料。     

ATP binding cassette transporters ABCG1 and ABCG16 affect reproductive development via auxin signalling in Arabidopsis

Angiosperm reproductive development is a complex event that includes floral organ development, male and female gametophyte formation and interaction between the male and female reproductive organs for successful fertilization. Previous studies have revealed the redundant function of ATP binding cassette subfamily G (ABCG) transporters ABCG1 and ABCG16 in pollen development, but whether they are involved in other reproductive processes is unknown. Here we show that ABCG1 and ABCG16 were not only expressed in anthers and stamen filaments but also enriched in pistil tissues, including the stigma, style, transmitting tract and ovule. We further demonstrated that pistil‐expressed ABCG1 and ABCG16 promoted rapid pollen tube growth through their effects on auxin distribution and auxin flow in the pistil. Moreover, disrupted auxin homeostasis in stamen filaments was associated with defective filament elongation. Our work reveals the key functions of ABCG1 and ABCG16 in reproductive development and provides clues for identifying ABCG1 and ABCG16 substrates in Arabidopsis.     

History of safe exposure and bioinformatic assessment of phosphomannose-isomerase (PMI) for allergenic risk

Newly expressed proteins in genetically engineered crops are evaluated for potential cross reactivity to known allergens as part of their safety assessment. This assessment uses a weight-of-evidence approach. Two key components of this allergenicity assessment include any history of safe human exposure to the protein and/or the source organism from which it was originally derived, and bioinformatic analysis identifying amino acid sequence relatedness to known allergens. Phosphomannose-isomerase (PMI) has been expressed in commercialized genetically engineered (GE) crops as a selectable marker since 2010 with no known reports of allergy, which supports a history of safe exposure, and GE events expressing the PMI protein have been approved globally based on expert safety analysis. Bioinformatic analyses identified an eight-amino-acid contiguous match between PMI and a frog parvalbumin allergen (CAC83047.1). While short amino acid matches have been shown to be a poor predictor of allergen cross reactivity, most regulatory bodies require such matches be assessed in support of the allergenicity risk assessment. Here, this match is shown to be of negligible risk of conferring cross reactivity with known allergens.