长爪沙鼠肝细胞体外分离培养体系的建立

目的建立长爪沙鼠原代肝细胞分离培养体系。方法以雄性长爪沙鼠为供体,采用组织消化法和Seglen两步灌流法分离肝细胞,以台盼蓝染色检测细胞得率和活率,过碘酸-希夫氏反应(PAS)鉴定肝细胞,倒置显微镜观察肝细胞形态变化,并使用含有多种细胞因子的培养基维持培养。结果组织消化法和Seglen两步灌流法平均每只长爪沙鼠可分别获得肝细胞(1.33±0.34)×107个、(3.97±1.15)×107个,细胞活率分别为(29.4±6.05)%、(80.3±4.56)%,这两种方法在细胞得率及活率方面存在显著差异。肝细胞内因有大量的糖原颗粒,经PAS染色后被染成红色。结果表明肝细胞在贴壁后72 h内,肝细胞形态发生显著变化。结论采用胶原酶经肝门静脉灌流分离肝细胞是一种高效获得肝细胞的方法。各种细胞因子有利于维持肝细胞在体外的生长分化,长爪沙鼠原代肝细胞分离培养体系的建立将为肝脏相关疾病研究和防治药物的开发提供技术支持。     

Glutamine residues in Q-loops of multidrug resistance protein MRP1 contribute to ATP binding via interaction with metal cofactor

Structural analyses of bacterial ATP-binding-cassette transporters revealed that the glutamine residue in Q-loop plays roles in interacting with: 1) a metal cofactor to participate in ATP binding; 2) a putative catalytic water molecule to participate in ATP hydrolysis; 3) other residues to transmit the conformational changes between nucleotide-binding-domains and transmembrane-domains, in ATP-dependent solute transport. We have mutated the glutamines at 713 and 1375 to asparagine, methionine or leucine to determine the functional roles of these residues in Q-loops of MRP1. All these single mutants significantly decreased Mg·ATP binding and increased the K(m) (Mg·ATP) and V(max) values in Mg·ATP-dependent leukotriene-C4 transport. However, the V(max) values of the double mutants Q713N/Q1375N, Q713M/Q1375M and Q713L/Q1375L were lower than that of wtMRP1, implying that the double mutants cannot efficiently bind Mg·ATP. Interestingly, MRP1 has higher affinity for Mn·ATP than for Mg·ATP and the Mn·ATP-dependent leukotriene-C4 transport activities of Q713N/Q1375N and Q713M/Q1375M are significantly higher than that of wtMRP1. All these results suggest that: 1) the glutamine residues in Q-loops contribute to ATP-binding via interaction with a metal cofactor; 2) it is most unlikely that these glutamine residues would play crucial roles in ATP hydrolysis and in transmitting the conformational changes between nucleotide-binding-domains and transmembrane-domains.     

The early life history of two sympatric New Zealand octopuses: eggs and paralarvae of Octopus huttoni and Pinnoctopus cordiformis

This study combined morphological and morphometric information on egg clutches, egg capsules and paralarvae of two sympatric coastal octopuses from New Zealand waters, Octopus huttoni and Pinnoctopus cordiformis, to provide species-specific traits to identify their early life stages obtained from field surveys. Eggs of O. huttoni (2.5 mm length; 1 mm width) were entwined with one another forming strings that ranged from 11 to 25.8 mm in length. Eggs of P. cordiformis (6.4 mm length; 1.5 mm width) were significantly bigger than those of O. huttoni and were grouped in small clusters of about seven eggs. Paralarvae O. huttoni and P. cordiformis differed in hatching size (1.4 mm versus 3.1 mm mantle length), number of suckers per arm (four versus eight), number of lamellae per outer demibranch (five versus ten) and arrangements of chromatophores in the body surface (29 to 59 versus 91 to 179), respectively. The morphological traits described in hatchlings from the laboratory allowed comparisons with field-collected paralarvae, suggesting that such characters were reliable species-specific patterns to enable a consistent differentiation between the early life stages of these two sympatric species, even in the absence of the brooding female.     

Effects of actinobacteria on plant disease suppression and growth promotion

Biological control and plant growth promotion by plant beneficial microbes has been viewed as an alternative to the use of chemical pesticides and fertilizers. Bacteria and fungi that are naturally associated with plants and have a beneficial effect on plant growth by the alleviation of biotic and abiotic stresses were isolated and developed into biocontrol (BCA) and plant growth-promoting agents (PGPA). Actinobacteria are a group of important plant-associated spore-forming bacteria, which have been studied for their biocontrol, plant growth promotion, and interaction with plants. This review summarizes the effects of actinobacteria as BCA, PGPA, and its beneficial associations with plants.     

Promoter-trapping in Saccharomyces cerevisiae by radiation-assisted fragment insertion

Non-homologous insertion (NHI) of DNA fragments into genomic DNA is a method widely used in insertional mutagenesis screens. In the yeast Saccharomyces cerevisiae, the efficiency of NHI is very low. Here we report that its efficiency can be increased by γ-irradiation of recipient cells at the time of transformation. Radiation-assisted NHI depends on YKU70, but its efficiency is not improved by inactivation of RAD5 or RAD52. In a pilot study, we generated 102 transformant clones expressing a lacZ reporter gene under standard conditions (30°C, rich medium). The site of insertion was determined in a subset of eight clones in which lacZ expression was altered by UV-irradiation. A comparison with published data revealed that three of the eight genes identified in our screen have not been targeted by large-scale transposon-based insertion screens. This suggests that radiation-assisted NHI offers a more homogeneous coverage of the genome than methods relying on transposons or retroviral elements.     

High throughput screening of hydrolytic enzymes from termites using a natural substrate derived from sugarcane bagasse

Background

Short rotation coppice willow is a potential lignocellulosic feedstock in the United Kingdom and elsewhere; however, research on optimising willow specifically for bioethanol production has started developing only recently. We have used the feedstock Salix viminalis × Salix schwerinii cultivar 'Olof' in a three-month pot experiment with the aim of modifying cell wall composition and structure within the stem to the benefit of bioethanol production. Trees were treated for 26 or 43 days with tension wood induction and/or with an application of the cellulose synthesis inhibitor 2,6-dichlorobenzonitrile that is specific to secondary cell walls. Reaction wood (tension and opposite wood) was isolated from material that had received the 43-day tension wood induction treatment.

Results

Glucan content, lignin content and enzymatically released glucose were assayed. All measured parameters were altered without loss of total stem biomass yield, indicating that enzymatic saccharification yield can be enhanced by both alterations to cell wall structure and alterations to absolute contents of either glucan or lignin.

Conclusions

Final glucose yields can be improved by the induction of tension wood without a detrimental impact on biomass yield. The increase in glucan accessibility to cell wall degrading enzymes could help contribute to reducing the energy and environmental impacts of the lignocellulosic bioethanol production process.     

实验动物皮肤病原真菌检测方法的改良

目的对实验动物皮肤病原真菌2种培养方法进行了比较。方法将采集到的3只皮肤真菌感染病兔样品经由沙氏平皿法和沙氏试管斜面培养法分别进行培养。结果在3只真菌感染病兔中应用试管斜面法我们只检测到1例皮肤病原真菌阳性,而采用沙氏平皿法3例阳性全部检出。结论结合临床检测经验,我们认为本研究的沙氏平皿法优于沙氏试管斜面法,在实验动物皮肤病原真菌常规检测中具有推广应用价值。     

羌活与宽叶羌活的群体遗传结构和种间分化研究

为了合理利用羌活和宽叶羌活的药用植物资源,同时保护其物种多样性,该研究利用SSR分子标记技术对羌活与宽叶羌活邻域及异域分布的23个自然种群,共计227个个体进行多样性和种间分化研究.结果显示:(1)两个物种具有中等水平的遗传多样性;羌活的平均等位基因数(Na)、有效等位基因数(Ne)和期望杂合度(He)分别为2.603...