Establishment of an in vitro culture system for chicken preblastodermal cells

To develop an alternative source for chicken pluripotent cells, we examined (1) whether undifferentiated preblastodermal cells could be subcultured in vitro for an extended period and (2) how subculturing affected the physiological properties of preblastodermal cells. The average number of preblastodermal cells was 2,397 in stage V embryos and 36,345 in stage VII embryos; stage X embryos had an average of 53,857 blastodermal cells. The average cell size decreased significantly (70.63-18.83 microm in diameter; P < 0.0001) as the embryo grew; this was closely related to a reduction in the size and number of lipid vesicles in the cell cytoplasm. The culture conditions were optimized for the stage V preblastodermal cells and the control stage X blastodermal cells. On STO feeder cells, the preblastodermal cells achieved stable growth in vitro only in HES medium or a mixed medium of the Knockout DMEM and HES media. However, more than 10 passages of preblastodermal cells at intervals of 3-4 days was possible only by using the Knockout/HES mixed medium and BRL cell-conditioned HES medium for the primary cultures and subcultures, respectively. Colony-forming preblastodermal cells had well-delineated cytoplasm, which was positively stained for stem cell-specific markers by anti-stage-specific embryo antigen-1 antibody, periodic acid-Schiff's solution, and alkaline phosphatase. When preblastodermal cells with or without culturing were transferred into the blastodermal cavity of stage X embryos, only in vitro-cultured preblastodermal cells at stage V (4/5 = 80%) and stage VII (2/8 = 25%) induced somatic chimerism in recipient chickens. In conclusion, undifferentiated preblastodermal cells could be subcultured, and only the colony-forming preblastodermal cells that stained positively for stem cell markers could induce somatic chimerism.     

婴幼儿配方奶粉菌落总数检验中不确定度的评定

分析样品中菌落总数不确定度的来源,采用合并样品标准差的方法来评定菌落总数的不确定。研究表明：实验中的扩展不确定度为0．046％,采用此法适合于菌落总数检验结果的评定。且该法简便,适合于每一个样本的检验结果,随着检验结果的不断增加,可随时加入到合并样本中,重新计算合并样本标准差,更新不确定度的取值范围。     

过表达外源M-CSF促进MCF7细胞增殖

为探讨过表达外源巨噬细胞集落刺激因子(M-CSF)对细胞增殖的影响,将重组载体 pCMV/cyto/myc-M-CSF转染MCF7细胞、G418筛选,RT-PCR、Western 印迹和免疫荧光鉴定M-CSF的mRNA表达、蛋白表达及定位,通过计算细胞倍增时间、MTT法及反义寡核苷酸抑制实验分析M-CSF对细胞增殖的影响. 结果表明,转染M-CSF的MCF7细胞过表达M-CSF-mRNA及蛋白,并且定位表达于细胞质;与转染空载体及未转染M-CSF组细胞比较,转染M-CSF的MCF7细胞倍增时间明显缩短、增殖能力显著增强,M-CSF的特异性反义寡核苷酸能抑制转染M-CSF的MCF7细胞的增殖,且抑制率随着反义寡核苷酸浓度的增高而增强;以上结果提示,胞质过表达M-CSF可促进MCF7细胞的增殖.     

用疏水色谱法复性并同时纯化重组人粒细胞-巨噬细胞集落刺激因子

目的：建立一种简单、快速复性并同时纯化大肠杆菌表达的重组人粒细胞一巨噬细胞集落刺激因子（rhGM-CSF）的方法。方法：研究rhGM-CSF在疏水色谱（HIC）上的复性和纯化机理,并对固定相和流动相进行选择和优化,包括固定相配基、流动相中盐的种类、流动相pH值、流动相中还原型谷胱甘肽（GSH）和氧化型谷胱甘肽（GSSG）的比例,以及流动相中尿素的浓度。结果：优化后的固定相为PEG600,流动相中的盐为（NH4）2SO4,流动相pH值为7．0,流动相中添加2．0mol／L尿素、1．8mmol／LGSH和0-3mmol／LGSSG。在优化条件下,HIC可使rhGM-CSF在分离纯化的同时得到复性,比活达1．58×10^7U／mg,纯度为95．7％,质量回收率为56．8％。结论：建立的疏水色谱复性和纯化工艺可简化操作步骤,缩短生产周期。     

Species cohesion despite extreme inbreeding in a social spider

Colonial social spiders experience extreme inbreeding and highly restricted gene flow between colonies; processes that question the genetic cohesion of geographically separated populations and which could imply multiple origins from predecessors with limited gene flow. We analysed species cohesion and the potential for long-distance dispersal in the social spider Stegodyphus dumicola by studying colony structure in eastern South Africa and the cohesion between this population and Namibian populations previously published. Data from both areas were (re)analysed for historic demographic parameters. Eastern South African S. dumicola were closely related to an east Namibian lineage, showing cohesion of S. dumicola relative to its sister species. Colony structure was similar in both areas with mostly monomorphic colonies, but haplotype diversity was much reduced in eastern South Africa. Here, the population structure indicated recent population expansion. By contrast, Namibia constitutes an old population, possibly the geographic origin of the species. Both the comparison of the eastern South African and Namibian lineages and the distribution within eastern South Africa show the potential for long-distance dispersal in few generations via colony propagation.     

G-protein-coupled receptor independent, immunomodulatory properties of chemokine CXCL9

Certain chemokines possess anti-angiogenic and antibacterial activity, in addition to their ability to recruit leukocytes. Herein, we demonstrate that CXCL9/MIG induces the expression, by a monocytic cell line and peripheral blood mononuclear cells, of a variety of chemokines including CXCL8/IL-8, CCL3/MIP-1α, CCL4/MIP-1β, CCL2/MCP-1 in a pertussis toxin insensitive manner. Similarly, another cationic chemokine CCL20/MIP-3α, but not the non-cationic chemokines CCL2 or CCL3, stimulated monocytic cells to produce substantial amounts of CXCL8 and CCL3. Microarray experiments demonstrated that CXCL9, but not CCL2, induced the expression of hundreds of genes, many of which have known or proposed immunomodulatory functions. Induction of CXCL8 required the p38 and ERK1/2 mitogen-activated protein kinases but not NFκB, JAK-STAT or JNK signaling pathways. These results collectively demonstrate that CXCL9 has immunomodulatory functions that are not mediated through a G-protein coupled receptor and may possess additional roles in host defenses against infection.     

Recruitment of Total Phytoplankton,Chlorophytes and Cyanobacteria from Lake Sediments Recorded by Photosynthetic Pigments in a Large,Shallow Lake (Lake Taihu,China)

Recruitment of total phytoplankton, chlorophytes and cyanobacteria from lake sediments to the water column was studied using photosynthetic pigments at one site (1.5 m) in Lake Taihu, a large shallow lake in China. Samples were taken weekly from the migration traps installed on the bottom from March to May 2004. Abundance of total phytoplankton, chlorophytes and cyanobacteria were represented by Chlorophyll (Chl) a, b, and phycocyanin (PC), respectively. Over the three months, total phytoplankton, chlorophytes, and cyanobacteria corresponding to 48.9%, 68.9% and 316.2% of their initial concentrations in surface sediments were recruited in Lake Taihu. However, compared with their increase in pelagic abundance over the same period, the recruitment accounted for a rather small inoculum. Accompanying the recruitment, total phytoplankton and chlorophytes declined and cyanobacteria increased in the upper 0–2 cm sediments; colonies of Microcystis aeruginosa in the water column enlarged from small size with several cells to large colonies with hundreds of cells. Thus, overwintering and subsequent growth renewal of pelagic phytoplankton merits further study and comparison with benthic survival and recruitment. (© 2005 WILEY‐VCH Verlag GmbH & Co. KGaA, Weinheim)     

基因工程菌生产rhG-CSF发酵培养基的研究

在摇瓶中对工程菌DH5αpBV220生产rhGCSF的发酵培养基进行了研究,从几种常用的细菌培养基中筛选适合工程菌表达rhGCSF的M9培养基。利用正交试验优化出生产重组人rhGCSF的优化培养基配方,其中优化碳源含量为2%的葡萄糖,氮源为25%的酵母粉、3%的蛋白胨和01%的(NH4)2SO4以及少量的无机盐。使用优化培养基可使rhGCSF的表达量达到338%,比优化前提高了489%。     

A sensitive, viable-colony staining method using Nile red for direct screening of bacteria that accumulate polyhydroxyalkanoic acids and other lipid storage compounds

The oxazine dye Nile blue A and its fluorescent oxazone form, Nile red, were used to develop a simple and highly sensitive staining method to detect poly(3-hydroxybutyric acid) and other polyhydroxyalkanoic acids (PHAs) directly in growing bacterial colonies. In contrast to previously described methods, these dyes were directly included in the medium at concentrations of only 0.5 μg/ml, and growth of the cells occurred in the presence of the dyes. This allowed an estimation of the presence of PHAs in viable colonies at any time during the growth experiment and a powerful discrimination between PHA-negative and PHA-positive strains. The presence of Nile red or Nile blue A did not affect growth of the bacteria. This viable-colony staining method was in particular applicable to gram-negative bacteria such as Azotobacter vinelandii, Escherichia coli, Pseudomonas putida, and Ralstonia eutropha. It was less suitable for discriminating between PHA-negative and PHA-positive strains of gram-positive bacteria such as Bacillus megaterium or Rhodococcus ruber, but it could also be used to discriminate between wax-ester- and triacylglycerol-negative and -positive strains of Acinetobacter calcoaceticus or Rhodococcus opacus. The potential of this new method and its application to further investigations of PHA synthases and PHA biosynthesis pathways are discussed. Received: 12 August 1998 / Accepted: 11 November 1998     

中国红光熊蜂(Bombus ignitus Smith)的饲养与观察

通过室内外饲养 ,观察了我国红光熊蜂蜂群的建立和发展动态。结果表明 ,在温度 2 8± 0 .5℃ ,相对湿度 70± 5 %和无光照的环境条件下 ,平均 84 .85 %越冬红光熊蜂蜂王在饲养后 11.83± 6 .13天开始产卵 ;加入欧洲熊蜂的幼工蜂能够刺激蜂王产卵 ,促使产卵期明显提前 ;健康且已受精的越冬蜂王一般在 2周内产卵 ;在开始饲养后 1个月左右开始出现工蜂 ,2月左右开始产生雄峰 ,雄峰出现后 7d左右产生蜂王。在采集并饲养的蜂王中 ,平均 78.79%的蜂王建立了蜂群 ,每群平均有蜂 180 .2 9± 80 .2 6头 ,其中工蜂 10 5 .90± 4 1.83头 ,雄蜂 6 5 .5 2± 4 5 .33头 ,蜂王 8.86± 12 .38头