Optimisation of overlap extension PCR-based mutagenesis of a GC-rich DNA template: application to the human α2C-adrenoceptor cDNA

PCR-based overlap extension mutagenesis was applied to introduce a Thr³⁸¹ to Lys mutation in the _2C-adrenoceptor (_2C AR) coding sequence. This cDNA contains 71% G+C nucleotides and conventional PCR procedures were inefficient in generating a corresponding amplification product. Only the combined use of a pre-PCR denaturation step at 100 °C followed by quick chilling on ice and the addition of 1 M of a commercial GC Melt reagent and 5% (v/v) dimethylsulphoxide with the Advantage GC cDNA PCR kit yielded efficient amplification during the three successive PCR steps of the overlap extension procedure. Transient expression of the mutant Thr³⁸¹Lys _2C AR in Cos-7 cells was performed to determine the binding profile for a series of ₂ AR ligands using [³H]RX 821002.     

藻类批量培养中的比增长率最大值

批量培养谷皮菱形藻、莱茵衣藻、粉核小球藻、斜生栅藻、羊角月牙藻和铜绿微囊藻等六种藻类,在对数生长期,逐日计数而确定比增长率最大值（μｍａｒ）,发现（μｍａｒ）值出现的时间在各种藻类中不同。同时讨论了每种藻类的（μｍａｒ）与营养物质、光强、温度和起始浓度之间的关系。铜绿微囊藻的（μｍａｒ）在一定的总磷（ＴＰ）浓度范围内随ＴＰ浓度呈上升趋势。根据获得增大率最大值的条件,对它们在浮游植物群落中的生态位作     

乌鲁木齐达坂城山区地衣生态位特征的研究

为揭示达坂城山区地衣对生境资源的利用状况及竞争程度,该研究在野外设置30个样方并调查其物种盖度数据,利用生态位宽度、生态位重叠指数及排序分析,探讨达坂城山区地衣的生态位特征及环境影响因子。结果表明:(1)达坂城山区生态位最宽的为丽黄鳞衣(Rusavskia elegans)和内卷野粮衣(Circinaria contorta),对不同环境具有较强的适应性,其他地衣物种生态位宽度较窄,对环境资源利用程度低。(2)物种间生态位重叠值整体偏低,生态位重叠值较高的物种对极少,地衣物种生态位分化程度高,物种间竞争不激烈。(3)生态位较宽的物种与多数物种之间都存在生态位重叠,但重叠值较低; 一些生态位较窄的物种与其他物种之间的生态位重叠值较高; 生态位重叠和生态位宽度之间并无直接的线性关系。(4)地衣物种沿海拔梯度分布具有差异,海拔、光照强度、湿度、风速和人为干扰是本地区地衣群落物种生态位特征差异的主导环境因子。综上所述,达坂城山区地衣物种由于生境资源的竞争占据各自独特的生态位,形成了生态位的分化,群落较为稳定,沿海拔梯度的生境条件差异是物种差异分布的重要原因。该研究可为地衣群落构建研究提供理论依据,并对该地区地衣物种的多样性和生境保护具有重要意义。     

Roles of endothermy in niche differentiation for ball-rolling dung beetles (Coleoptera: Scarabaeidae) along an altitudinal gradient

Abstract.  1. An analysis of whether niche differentiation in ball-rolling dung beetles can be explained by the way in which they regulate their body temperature was conducted.
2.  A priori assumptions were: (i) if thermoregulation affects niche partitioning, sympatric species must have different endothermic strategies that minimise encounters; or, alternatively (ii) if two co-occurring species show the same thermoregulation pattern and their flight periods overlap, they might be avoiding competition by exhibiting different resource preferences or different food relocation behaviour.
3. The ball-rolling dung beetles studied showed a hierarchical structure based on the species' endothermic capacity, measured as temperature excess [ T _ex= difference between body temperature ( T _b) and ambient temperature ( T _a)]. Those with a high T _ex (10–15 °C) were located exclusively at altitudes >1000 m a.s.l. On the coastal plains, species with a high T _ex were restricted to flying at night when the T _a was lower. Species with a lower T _ex (less than 10 °C higher than T _a) were found in the coastal plains zone.
4. Where there was sympatry with similar trophic habits, the species involved showed very different thermal niches, and where there was significant overlap of thermal niches between sympatric species, trophic habits of species were very different.
5. The results suggest that it is possible to use the concept of the thermal niche as a tool to explain interspecific interactions and the spatial distribution of species.     

Relationships between bird morphology and prey selection in two sympatric Great Cormorant Phalacrocorax carbo subspecies during winter

Variation in bird morphology (notably sex size dimorphism) has been suggested to contribute to differences in food use between individuals. We explore the hypothesis of food partitioning (diet overlap and prey size selection) in two sympatric subspecies of the Great Cormorant Phalacrocorax carbo with respect to bird morphology (subspecies and sex) in inland French waters. These areas represent a recent and increasing contact zone used as a common non-territorial winter feeding area by the continental P. c. sinensis and the marine P. c. carbo subspecies. A high dietary overlap between subspecies and sexes was found at the major sites studied. Prey size selection was found to be site-specific and generally related to a gradient of structural size and body mass (male P. c. carbo  > male P. c. sinensis  > female P. c. carbo  > female P. c. sinensis ). With respect to bird morphology, differences in prey size consumed by Great Cormorants were more pronounced between sexes in freshwater habitats than between subspecies. This was reinforced by the fact that P. c. carbo birds entering inland areas were smaller than birds on the coast. These results also suggest that bird morphology is an important determinant of dietary differences. Our study demonstrated that P. c. carbo is an efficient generalist forager in continental areas, and food partitioning in type and size of prey between the two subspecies is low.     

Discrimination of grasshopper (Orthoptera: Acrididae) diet and niche overlap using next‐generation sequencing of gut contents

Species of grasshopper have been divided into three diet classifications based on mandible morphology: forbivorous (specialist on forbs), graminivorous (specialist on grasses), and mixed feeding (broad‐scale generalists). For example, Melanoplus bivittatus and Dissosteira carolina are presumed to be broad‐scale generalists, Chortophaga viridifasciata is a specialist on grasses, and Melanoplus femurrubrum is a specialist on forbs. These classifications, however, have not been verified in the wild. Multiple specimens of these four species were collected, and diet analysis was performed using DNA metabarcoding of the gut contents. The rbcLa gene region was amplified and sequenced using Illumina MiSeq sequencing. Levins' measure and the Shannon–Wiener measure of niche breadth were calculated using family‐level identifications and Morisita's measure of niche overlap was calculated using operational taxonomic units (OTUs). Gut contents confirm both D. carolina and M. bivittatus as generalists and C. viridifasciata as a specialist on grasses. For M. femurrubrum, a high niche breadth was observed and species of grasses were identified in the gut as well as forbs. Niche overlap values did not follow predicted patterns, however, the low values suggest low competition between these species.     

Unique biomechanical interactions between myeloma cells and bone marrow stroma cells

We observed that BMSCs (bone marrow stromal cells) from myeloma patients (myeloma BMSCs) were significantly stiffer than control BMSCs using a cytocompression device. The stiffness of myeloma BMSCs and control BMSCs was further increased upon priming by myeloma cells. Additionally, myeloma cells became stiffer when primed by myeloma BMSCs. The focal adhesion kinase activity of myeloma cells was increased when cells were on stiffer collagen gels and on myeloma BMSCs. This change in myeloma stiffness is associated with increased colony formation of myeloma cells and FAK activation when co-cultured with stiffer myeloma BMSCs or stiffer collagen. Additionally, stem cells of RPMI8226 cells became stiffer after priming by myeloma BMSCs, with concomitant increases of stem cell colony formation. These results suggest the presence of a mechanotransduction loop between myeloma cells and myeloma BMSCs to increase the stiffness of both types of cells via FAK activation. The increase of stiffness may in turn support the growth of myeloma cells and myeloma stem cells.     

Human CD133-derived bone marrow stromal cells establish ectopic hematopoietic microenvironments in immunodeficient mice

Cultured adherent bone marrow stromal cells (BMSCs) are capable of forming ectopic hematopoietic microenvironments (HMEs) in immunodeficient mice. However, the cell surface phenotype of the native bone marrow stem/progenitor cell that gives rise to BMSCs that support hematopoiesis remains poorly defined. We recently reported the derivation of human BMSC-like cells (CD133BMSCs) by magnetic cell sorting against Prominin-1 (CD133), an epitope expressed by embryonic, fetal, and adult stem cells. Here we demonstrate that CD133BMSCs are capable of forming ectopic HMEs. Cultured adherent CD133BMSCs derived from sorted CD133-positive cells lacked CD133 expression, but were uniformly positive for CD146, an epitope recently described to identify self-renewing osteoprogenitor cells that could transfer the HME. CD133BMSCs were genetically-tagged by lentivirus, expanded, and seeded into HA/TCP/fibrin constructs that were implanted subcutaneously. After 60 days, CD133BMSCs produced human osteocytes, osteoblasts, adipocytes, and reticular cells that supported murine hematopoiesis. CD133BMSCs that were not transduced with lentivirus also formed HMEs. Control constructs seeded with human dermal fibroblasts formed connective tissue, but failed to form HMEs. Our data indicate that CD133 expression identifies a native human bone marrow stem/progenitor cell that gives rise to BMSCs capable of forming the HME.     

Niche diversity in crustacean cryptic species: complementarity in spatial distribution and predation risk

Recent genetic studies indicate that species with very close phenotypic similarity (“cryptic species”) are a common feature of nature, and that such cryptic species often coexist in communities. Because traditional views of species coexistence demand that species differ in phenotype to coexist stably, the existence of sympatric cryptic species appears to challenge traditional perspectives of coexistence. We evaluated niche diversity in three recently discovered species of Hyalella amphipods that occur sympatrically in lakes and share close phenotypic similarity. We found that, in some cases, these species exhibited strong complementary spatial distributions within the littoral zone of lakes, both across a distance-from-shore gradient, and a vertical depth gradient. Additionally, we compared fish stomach contents with habitat samples and found that species differed in their vulnerability to predation from sunfish (Lepomis spp.). Complementarity among species across axes of spatial distribution and predation risk, two important niche components, suggests that species with close phenotypic similarity may differ appreciably along ecologically relevant axes. Our results, considered in the light of previous studies, suggest a community structured by predator-mediated coexistence or sequential dominance across environmental gradients in the littoral zone.