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Image Credit: BioProtoc.2669

生物化学

Determining Ribosome Translational Status by Ribo-ELISA 采用Ribo-ELISA检测核糖体的翻译状态 作者:Amandine Bastide, Jonathan W. Yewdell and Alexandre David日期:01/05/2018,浏览量:5136,Q&A: 0
The Ribo-ELISA was originally developed to elucidate the basis for the ribopuromycylation method (RPM)-based detection of ribosome bound nascent chains. The Ribo-ELISA enables characterization of the translational status of ribosomes, and can be applied to the discovery of super-ribosomal complexes with novel ribosome associated macromolecules ...
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细胞生物学

High Throughput NPY-Venus and Serotonin Secretion Assays for Regulated Exocytosis in Neuroendocrine Cells 高通量NPY-Venus和5-羟色胺分泌实验分析神经内分泌细胞中被调控的胞吐作用 作者:Xingmin Aaron Zhang and Thomas F.J. Martin日期:01/05/2018,浏览量:5102,Q&A: 0
Here we describe two assays to measure dense core vesicle (DCV) exocytosis-mediated cargo secretion in neuroendocrine cells. To conduct siRNA screens for novel genes in regulated DCV exocytosis, we developed a plate reader-based secretion assay using DCV cargo, NPY-Venus, and an orthogonal 3H-serotonin secretion assay. The NPY-Venus ...
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免疫学

Detection of Intracellular Reduced (Catalytically Active) SHP-1 and Analyses of Catalytically Inactive SHP-1 after Oxidation by Pervanadate or H2O2 细胞内还原的(具有催化活性的)SHP-1检测以及在经钒酸盐或H2O2氧化后无催化活性SHP-1的分析 作者:Seeyoung Choi and Paul E. Love日期:01/05/2018,浏览量:4961,Q&A: 0
Oxidative inactivation of cysteine-dependent Protein Tyrosine Phosphatases (PTPs) by cellular reactive oxygen species (ROS) plays a critical role in regulating signal transduction in multiple cell types. The phosphatase activity of most PTPs depends upon a ‘signature’ cysteine residue within the catalytic domain that is maintained in the ...
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微生物学

Easy and Efficient Permeabilization of Cyanobacteria for in vivo Enzyme Assays Using B-PER 使用B-PER简单有效地透化蓝藻以进行体内酶分析 作者:Simon Matthé Erstad and Yumiko Sakuragi日期:01/05/2018,浏览量:5532,Q&A: 0
Cyanobacteria are photosynthetic bacteria that thrive in diverse ecosystems and play major roles in the global carbon cycle. The abilities of cyanobacteria to fix atmospheric CO2 and to allocate the fixed carbons to chemicals and biofuels have attracted growing attentions as sustainable microbial cell factories. A better understanding ...
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Targeted Genome Editing of Virulent Phages Using CRISPR-Cas9 使用CRISPR-Cas9对烈性噬菌体进行靶向基因组编辑 作者:Marie-Laurence Lemay, Ariane C. Renaud, Geneviève M. Rousseau and Sylvain Moineau日期:01/05/2018,浏览量:12904,Q&A: 1
This protocol describes a straightforward method to generate specific mutations in the genome of strictly lytic phages. Briefly, a targeting CRISPR-Cas9 system and a repair template suited for homologous recombination are provided inside a bacterial host, here the Gram-positive model Lactococcus lactis MG1363. The CRISPR-Cas9 system is ...
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Analysis of Direct Interaction between Viral DNA-binding Proteins by Protein Pull-down Co-immunoprecipitation Assay 蛋白质pull-down免疫共沉淀实验分析病毒DNA结合蛋白质之间的直接相互作用 作者:Ana Lechuga, Mónica Berjón-Otero, Margarita Salas and Modesto Redrejo-Rodríguez日期:01/05/2018,浏览量:7928,Q&A: 0
This protocol analyzes the direct interaction between two DNA-binding proteins by pull-down co-immunoprecipitation. One of the proteins is overexpressed in E. coli as HA-tagged recombinant protein and cell-free extracts are immunoprecipitated in HA-affinity resin. Cell extracts are treated with nuclease to degrade DNA and RNA, which rules ...
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Transplantation of Fecal Microbiota Shaped by Diet 饮食经控制的粪便微生物群的移植 作者:Keiran H. McLeod, Linda Mason and Eliana Mariño日期:01/05/2018,浏览量:7119,Q&A: 0
Alterations in diet and gut microbial ecology underlie the pathogenesis of type 1 diabetes (T1D). In the non-obese diabetic (NOD) mouse, we found high concentrations of bacterial metabolites acetate and butyrate in blood and faeces correlated with protection from disease. We reconstituted germ free (GF) NOD mice with fecal bacteria from protected ...
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Bacterial Aggregation Assay in the Presence of Cyclic Lipopeptides 环脂肽存在情况下细菌聚集实验 作者:Pengyuan Xiu, Rui Liu, Dechao Zhang and Chaomin Sun日期:01/05/2018,浏览量:6634,Q&A: 0
Lipopeptides is an important class of biosurfactants having antimicrobial and anti-adhesive activity against pathogenic bacteria. These include surfactin, fengycin, iturin, bacillomycin, mycosubtilin, lichenysin, and pumilacidin (Arima et al., 1968; Naruse et al., 1990; Yakimov et al., 1995; Steller and Vater, 2000; ...
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神经科学

A Rodent Model for Chronic Brain Hypoperfusion Related Diseases: Permanent Bilateral Occlusion of the Common Carotid Arteries (2VO) in Rats 慢性脑灌注不足相关疾病的啮齿动物模型:大鼠双侧颈总动脉永久性闭塞(2VO) 作者:Mei-Ling Yan and Jing Ai日期:01/05/2018,浏览量:6973,Q&A: 0
Permanent occlusion of bilateral common carotid arteries (2VO) in rat is considered as a suitable animal model to mimic chronic brain hypoperfusion status, which is proved to be a risk factor to precede the Alzheimer’s disease or/and vascular dementia. In this protocol, we describe how to successfully ligate the bilateral common carotid arteries ...
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The RiboPuromycylation Method (RPM): an Immunofluorescence Technique to Map Translation Sites at the Sub-cellular Level RiboPuromycylation方法(RPM):一种在亚细胞水平绘制翻译位点图谱的免疫荧光技术 作者:Amandine Bastide, Jonathan W. Yewdell and Alexandre David日期:01/05/2018,浏览量:10051,Q&A: 0
While isotopic labeling of amino acids remains the reference method in the field for quantifying translation rate, it does not provide any information on spatial localization of translation sites. The rationale behind developing the ribopuromycylation method (RPM) was primarily to map translation sites at the sub-cellular level while avoiding ...
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Mutant Huntingtin Secretion in Neuro2A Cells and Rat Primary Cortical Neurons Neuro2A细胞和大鼠原代皮层神经元中突变亨廷顿蛋白的分泌 作者:Katarina Trajkovic, Hyunkyung Jeong and Dimitri Krainc日期:01/05/2018,浏览量:5621,Q&A: 0
Quantitative analysis of proteins secreted from the cells poses a challenge due to their low abundance and the interfering presence of a large amount of bovine serum albumin (BSA) in the cell culture media. We established assays for detection of mutant huntingtin (mHtt) secreted from Neuro2A cell line stably expressing mHtt and rat primary ...
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Behavioral Assays to Study Oxygen and Carbon Dioxide Sensing in Caenorhabditis elegans 研究秀丽隐杆线虫的氧气和二氧化碳传感的行为分析实验 作者:Teresa Rojo Romanos, Leelee Ng, Manuel Zimmer and Roger Pocock日期:01/05/2018,浏览量:5263,Q&A: 0
Animals use behavioral strategies to seek optimal environments. Population behavioral assays provide a robust means to determine the effect of genetic perturbations on the ability of animals to sense and respond to changes in the environment. Here, we describe a C. elegans population behavioral assay used to measure locomotory responses ...
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Visible Immunoprecipitation (VIP) Assay: a Simple and Versatile Method for Visual Detection of Protein-protein Interactions 可视免疫沉淀(VIP)实验:一种简单通用的蛋白质 - 蛋白质相互作用可视化检测方法 作者:Yohei Katoh, Kentaro Nakamura and Kazuhisa Nakayama日期:01/05/2018,浏览量:10016,Q&A: 0
The visible immunoprecipitation (VIP) assay is a convenient alternative to conventional co-immunoprecipitation (Katoh et al., 2015). By processing lysates from cells co-expressing GFP-fusion and RFP-fusion proteins for immunoprecipitation with GST-tagged anti-GFP Nanobody and glutathione-Sepharose beads, protein-protein interactions can ...
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植物科学

Rolling Circle Amplification to Screen Yam Germplasm for Badnavirus Infections and to Amplify and Characterise Novel Badnavirus Genomes 滚换扩增筛选山药Badnavirus感染种质并扩增鉴定新型Badnavirus基因组 作者:Moritz Bömer, Aliyu A. Turaki, Ajith I. Rathnayake, Gonçalo Silva, P. Lava Kumar and Susan E. Seal日期:01/05/2018,浏览量:7698,Q&A: 0
Since the first discovery of badnaviruses (family Caulimoviridae, genus Badnavirus) in yam (Dioscorea spp.) germplasm in the 1970s (Harrison and Roberts, 1973), several hundred partial badnavirus reverse transcriptase (RT)-ribonuclease H (RNaseH) sequences have been characterised (Kenyon et al., 2008; Bousalem ...
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In vitro RNA-dependent RNA Polymerase Assay Using Arabidopsis RDR6 拟南芥RDR6的体外RNA依赖性RNA聚合酶活性测定 作者:Kyungmin Baeg, Yukihide Tomari and Hiro-oki Iwakawa日期:01/05/2018,浏览量:8008,Q&A: 0
RNA-dependent RNA polymerases (RdRPs) in eukaryotes convert single-stranded RNAs into double-stranded RNAs, thereby amplifying small interfering RNAs that play crucial roles in the regulation of development, maintenance of genome integrity and antiviral immunity. Here, we describe a method of in vitro RdRP assay using recombinant ...
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Fatty Acid Content and Composition of Triacylglycerols of Chlorella kessleri 凯氏小球藻中脂肪酸含量及三酰甘油组成 作者:Motohide Aoki and Norihiro Sato日期:01/05/2018,浏览量:6944,Q&A: 0
Triacylglycerols (TAGs) are esters formed from one glycerol and three fatty acids. TAGs are induced to accumulate in algal cells under environmental stress conditions including nutrient-limitation, hyperosmosis, and low temperature, for the storage of metabolic energy and carbon, and also for the consumption of excess energy (e.g., Hirai ...
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Chromatin Affinity Purification (ChAP) from Arabidopsis thaliana Rosette Leaves Using in vivo Biotinylation System 使用体内生物素化系统进行拟南芥莲座叶染色质亲和纯化(ChAP) 作者:Weronika Sura and Piotr A. Ziolkowski日期:01/05/2018,浏览量:6726,Q&A: 0
Chromatin Affinity Purification (ChAP) is widely used to study chromatin architecture and protein complexes interacting with DNA. Here we present an efficient method for ChAP from Arabidopsis thaliana rosette leaves, in which in vivo biotinylation system is used. The chromatin is digested by Micrococcal Nuclease (MNase), hence ...
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Electron Tomography to Study the Three-dimensional Structure of Plasmodesmata in Plant Tissues–from High Pressure Freezing Preparation to Ultrathin Section Collection 采用电子断层摄影术研究植物组织中的胞间连丝的三维结构 — 从高压冷冻制备到超薄切片采集 作者:William J. Nicolas, Emmanuelle Bayer and Lysiane Brocard日期:01/05/2018,浏览量:7444,Q&A: 0
Plasmodesmata (PD) are nanometric (~20 nm wide) membrane lined pores encased in the cell walls of the adjacent plant cells. They allow the cells to exchange all types of molecules ranging from nutrients like sugar, hormones, to RNAs and various proteins. Unfortunately, they are also hijacked by phyto-viruses, enabling them to spread from ...
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Micro-computed Tomography to Visualize Vascular Networks in Maize Stems 玉米茎杆中维管网络的微型计算机断层摄影术观察 作者:Akiteru Maeno and Katsutoshi Tsuda日期:01/05/2018,浏览量:7961,Q&A: 0
Plant vascular systems in the stem connect roots with aerial organs to move solutes containing minerals, nutrients as well as signaling molecules, and therefore, they play pivotal roles in plant growth and development. However, stem vascular systems, especially in crop species, have been poorly described since they are deeply embedded in the ...
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Real-time Analysis of Auxin Response, Cell Wall pH and Elongation in Arabidopsis thaliana Hypocotyls 拟南芥下胚轴植物生长素响应、细胞壁pH和伸长的实时分析 作者:Lanxin Li, S. F. Gabriel Krens, Matyáš Fendrych and Jiří Friml日期:01/05/2018,浏览量:7399,Q&A: 0
The rapid auxin-triggered growth of the Arabidopsis hypocotyls involves the nuclear TIR1/AFB-Aux/IAA signaling and is accompanied by acidification of the apoplast and cell walls (Fendrych et al., 2016). Here, we describe in detail the method for analysis of the elongation and the TIR1/AFB-Aux/IAA-dependent auxin response in ...
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