大白菜抗软腐病基因BrWRKY33植物表达载体的构建

利用农杆菌侵染法获得抗软腐病转BrWRKY33基因大白菜,首先要构建植物表达载体。本实验根据BrWRKY33基因的序列及pROK2表达载体的酶切位点设计引物(FN/RN),获得BrWRKY33基因,然后将该基因连接到克隆载体pGEM-TEasy,重新构建了克隆载体T-WRKY。经测序及酶切验证,选取PCR反应中错配率最低的产物,构建了表达载体pROK2-WRKY,并将该表达载体转入到根癌农杆菌EHA105中,并对其转化子进行了鉴定。结果表明,PCR扩增获得目的基因BrWRKY33全长约1443bp,PCR及酶切结果鉴定表明克隆载体T-WRKY已经重新构建成功。表达载体的PCR及BamHⅠ和KpnⅠ双酶切鉴定表明有4个重组子表现阳性,表明目的基因已经插入到pROK2表达载体,表达载体构建成功。PCR鉴定表明表达载体pROK2-WRKY已经转入根癌农杆菌EHA105中。本实验结果将为进一步研究大白菜抗软腐病基因的转化奠定基础。     

Binding of nonsteroidal anti‐inflammatory drugs to Aβ fibril

Nonsteroidal anti‐inflammatory drugs are considered as potential therapeutic agents against Alzheimer's disease. Using replica exchange molecular dynamics and atomistic implicit solvent model, we studied the mechanisms of binding of naproxen and ibuprofen to the Aβ fibril derived from solid‐state NMR measurements. The binding temperature of naproxen is found to be almost 40 K higher than of ibuprofen implicating higher binding affinity of naproxen. The key factor, which enhances naproxen binding, is strong interactions between ligands bound to the surface of the fibril. The naphthalene ring in naproxen appears to provide a dominant contribution to ligand‐ligand interactions. In contrast, ligand‐fibril interactions cannot explain differences in the binding affinities of naproxen and ibuprofen. The concave fibril edge with the groove is identified as the primary binding location for both ligands. We show that confinement of the ligands to the groove facilitates ligand‐ligand interactions that lowers the energy of the ligands bound to the concave edge compared with those bound to the convex edge. Our simulations appear to provide microscopic rationale for the differing binding affinities of naproxen and ibuprofen observed experimentally. Proteins 2010. © 2010 Wiley‐Liss, Inc.     

HIV-1 CN54 Pol P51抗原高效表达、纯化、复性及在抗体检测中的应用

为获得可溶性高纯度HIV-1中国株CN54PolP51抗原,将携带CN54polp51基因的重组质粒pTHioHisA51转化大肠杆菌BL21codonplus-RIL,用IPTG进行诱导表达。用Chelating SepharoseFF-Ni亲和层析柱及DEAE Sepharose Fast Flow阴离子交换层析柱纯化目的蛋白,采用透析复性法得到可溶性抗原,Western blotting检测目的蛋白。用纯化的P51抗原蛋白标记辣根过氧化物酶及包被酶标板进行双抗原夹心法ELISA检测。结果显示P51以包涵体的形式表达,表达量占菌体总蛋白的50%,经两步层析和透析复性,目的蛋白纯度大于95%。Western blotting和双抗原夹心法ELISA检测均显示了良好的灵敏度和特异性。本研究可以为HIV-1疫苗研究和开发检测试剂提供支持。     

Hollow Palladium‐Gold Nanochains with Periodic Concave Structures as Superior ORR Electrocatalysts and Highly Efficient SERS Substrates

Unique palladium‐gold hollow nanochains (PdAu HCs) with 1D architecture and an ultrathin Pd‐rich skin exhibit fantastic oxygen reduction reaction (ORR) enhancements due to their high conductivity, structural stability, and maximized atomic utilization. More importantly, such PdAu HCs possess periodic concave structures that boost ORR performance. These structures readily form high‐density high‐index facets, and fewer arc edges. Concave structures can deliver strong strain effects and surface charge accumulation is revealed by off‐axis electron holography. In addition, periodic concave structures can provide strong localized surface–plasmon coupling, which means that PdAu HCs have great potential as efficient surface‐enhanced Raman scattering (SERS) substrates. This study offers a novel and general approach for the design of complex noble metal‐based nanostructures as efficient ORR catalysts and SERS substrates.     

Age-dependent tree-ring growth responses of Schrenk spruce (Picea schrenkiana) to climate—A case study in the Tianshan Mountain,China

For both its climatic and ecological importance, Schrenk spruce (Picea schrenkiana) is a crucial tree species living at mid-altitude on the western area of the Tianshan Mountains. It plays a key role on understanding climatic change in the Tianshan Mountains in the past 500 years. However, whether the relationship between tree growth and limiting climate factors is stable over time is still not well-known. In this study, standard and residual chronologies of four 100-year age classes (AC1 < 110a, 110a < AC2 < 210a, 210a < AC3 < 310a and AC4 > 310a) were established for detecting divergence in climate–growth relationships as well as comparing low-frequency and high-frequency variations. The results show that climate can account for a high amount of variance in tree-ring width and higher climate sensitivity was detected in younger trees. Younger trees (<210a) exhibit significantly negative growth responses to mean monthly air temperature of previous June and positive relationship with total monthly precipitation of current April and May, while mean monthly air temperature of current March may inhibit growth of older trees (>210a). Tree-ring chronology statistics and response function reveal that the age-growth patterns are non-monotonic. Our results together with previous studies demonstrate that the age effects on tree-ring growth–climate response is attributed to a combination of genetic characteristics and site microclimate, which suggests that it is necessary to consider both age-dependent and species-specific climate responses when using tree-ring measurements as a proxy for valid climate reconstructions.     

CST-II's recognition domain for acceptor substrates in α-(2→8)-sialylations

CST-II is a bacterial sialyltransferase known for its ability to perform α-(2→8)-sialylations using GM(3) related trisaccharide substrates. Previously, we probed the enzyme's substrate specificity and developed an efficient synthesis for α-(2→8)-oligosialosides, and we suggested that CST-II could have a very small substrate recognition domain. Here we report our full studies on CST-II's recognition feature for acceptor substrates. The current study further demonstrates the versatility of CST-II in preparing complex oligosaccharides that contain α-(2→8)-oligosialyl moieties.     

PheMaDB: A solution for storage,retrieval, and analysis of high throughput phenotype data

Background  

OmniLog™ phenotype microarrays (PMs) have the capability to measure and compare the growth responses of biological samples upon exposure to hundreds of growth conditions such as different metabolites and antibiotics over a time course of hours to days. In order to manage the large amount of data produced from the OmniLog™ instrument, PheMaDB (Phenotype Microarray DataBase), a web-based relational database, was designed. PheMaDB enables efficient storage, retrieval and rapid analysis of the OmniLog™ PM data.     

Evaluation of the prognostic value of TGF-β superfamily type I receptor and TGF-β type II receptor expression in nasopharyngeal carcinoma using high-throughput tissue microarrays

Gene expression profiling had revealed that TGF-β superfamily type I receptor (also known as activin receptor-like kinase-1, ALK1) and TGFβR2 (TGF-β type II receptor) were down-regulated in nasopharyngeal carcinoma (NPC) (P < 0.05, respectively). However, no study with significantly large clinical samples to address the relevance of ALK1 and TGFβR2 in NPC progression or in patient outcomes has been reported. This study aims to assess the possible correlations of ALK1 and TGFβR2 expression with NPC progression and their potential prognostic predictive ability in NPC outcomes. ALK1 and TGFβR2 mRNA and protein levels were detected by qRT-PCR and NPC tissue microarray (TMA), which included 742 tissue cores. Both mRNA and protein levels of ALK1 and TGFβR2 were significantly lower in the cancer tissues compared with the non-cancerous tissues (P < 0.05). Epstein-Barr virus small RNA (EBER-1) hybridization signals in NPC showed significant associations with ALK1 and TGFβR2 proteins (P = 0.000 and 0.003, respectively). In the final logistic regression analysis model, the abnormal expression of ALK1 and TGFβR2 were found to be independent contributors to nasopharyngeal carcinogenesis (P = 0.000 and 0.000, respectively). A survival analysis revealed that ALK1 (Disease Free Survival (DFS): P = 0.002, Overall Survival (OS): P = 0.007) and TGFβR2 (DFS: P = 0.072, OS: P = 0.045) could predict the prognosis of NPC patients. The positive expression of ALK1 and TGFβR2 were independent risk factors for DFS and OS in multivariate analyses (DFS: P = 0.001 and 0.420, respectively; OS: P = 0.018 and 0.047, respectively). These results suggest that ALK1 and TGFβR2 may be useful prognostic biomarkers in NPC.