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Latest Articles

Investigating Protein Structure and Evolution with SCOP2
Antonina Andreeva, Dave Howorth, Cyrus Chothia, Eugene Kulesha, Alexey G. Murzin

SCOP2 is a successor to the Structural Classification of Proteins (SCOP) database that organizes proteins of known structure according to their structural and evolutionary relationships. It was designed to provide a more advanced framework for the classification of proteins.

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Scoring Large‐Scale Affinity Purification Mass Spectrometry Datasets with MiST
Erik Verschueren, John Von Dollen, Peter Cimermancic, Natali Gulbahce, Andrej Sali, Nevan J. Krogan

High‐throughput Affinity Purification Mass Spectrometry (AP‐MS) experiments can identify a large number of protein interactions, but only a fraction of these interactions are biologically relevant.

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Expression Data Analysis with Reactome
Steve Jupe, Antonio Fabregat, Henning Hermjakob

The Reactome database of curated biological pathways provides a tool for visualizing user‐supplied expression data as an overlay on pathway diagrams, thereby affording an effective means to examine expression of the constituents of the pathway and determine whether all that are necessary ar.

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Using pLink to Analyze Cross‐Linked Peptides
Sheng‐Bo Fan, Jia‐Ming Meng, Shan Lu, Kun Zhang, Hao Yang, Hao Chi, Rui‐Xiang Sun, Meng‐Qiu Dong, Si‐Min He

pLink is a search engine for high‐throughput identification of cross‐linked peptides from their tandem mass spectra, which is the data‐analysis step in chemical cross‐linking of proteins coupled with mass spectrometry analysis.

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Using REDItools to Detect RNA Editing Events in NGS Datasets
Ernesto Picardi, Anna Maria D'Erchia, Antonio Montalvo, Graziano Pesole

RNA editing is a post‐transcriptional/co‐transcriptional molecular phenomenon whereby a genetic message is modified from the corresponding DNA template by means of substitutions, insertions, and/or deletions.

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Basic Techniques in Mammalian Cell Tissue Culture
Katy Phelan, Kristin M. May

Cultured mammalian cells are used extensively in cell biology studies. It requires a number of special skills in order to be able to preserve the structure, function, behavior, and biology of the cells in culture.

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Isolation and Culture of Spinal Cord Motor Neurons
Alice Klausmeyer, D. Stern, S. Wiese

Isolated spinal motoneurons are a powerful tool for studying basic mechanisms of neurite growth and survival. Since motoneurons are a minor population of developing spinal cord cells, they need to be purified and enriched to separate them from non‐neuronal cells.

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Isolation of Integrin‐Based Adhesion Complexes
Matthew C. Jones, Jonathan D. Humphries, Adam Byron, Angélique Millon‐Frémillon, Joseph Robertson, Nikki R. Paul, Daniel H. J. Ng, Janet A. Askari, Martin J. Humphries

The integration of cells with their extracellular environment is facilitated by cell surface adhesion receptors, such as integrins, which play important roles in both normal development and the onset of pathologies.

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Adhesive Micropatterns to Study Intermediate Filament Function in Nuclear Positioning
Isabelle Dupin, Julien Elric, Sandrine Etienne‐Manneville

The nucleus is generally found near the cell center; however its position can vary in response to extracellular or intracellular signals, leading to a polarized intracellular organization. Nuclear movement is mediated by the cytoskeleton and its associated motors.

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Reconstitution of a Bioengineered Salivary Gland Using a Three‐Dimensional Cell Manipulation Method
Miho Ogawa, Takashi Tsuji

One concept in regenerative therapy is the replacement of a lost or damaged organ with a regenerated, fully functional organ.

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Diffraction‐Unlimited Fluorescence Microscopy of Living Biological Samples Using pcSOFI
Sam Duwé, Benjamien Moeyaert, Peter Dedecker

The complex microscopic nature of many live biological processes is often obscured by the diffraction limit of light, requiring diffraction‐unlimited fluorescence microscopy to resolve them.

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Preparation and Separation of DNA‐Wrapped Carbon Nanotubes
Geyou Ao, Ming Zheng

Purification of single‐chirality single‐wall carbon nanotubes (SWCNTs) from their synthetic mixture is a prerequisite for many applications.

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SNIPER Peptide‐Mediated Degradation of Endogenous Proteins
Xuelai Fan, Yu Tian Wang

Rapid and reversible methods for altering the function of endogenous proteins are not only indispensable tools for probing complex biological systems, but may potentially drive the development of new therapeutics for the treatment of human diseases.

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Universal Non‐Antibody Detection of Protein Phosphorylation Using pIMAGO
Anton B. Iliuk, W. Andy Tao

This article describes methods for a new, non‐antibody phosphorylation detection reagent, termed pIMAGO (phospho‐imaging).

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Glucose Tolerance Tests for Systematic Screening of Glucose Homeostasis in Mice
Jan Rozman, Birgit Rathkolb, Susanne Neschen, Helmut Fuchs, Valérie Gailus‐Durner, Martin Klingenspor, Eckhard Wolf, Martin Hrabě de Angelis

This article presents a detailed description of intraperitoneal and oral glucose tolerance tests in mice. The former is widely used in initial high‐throughput phenotyping of mutant mice to assess a diabetic phenotype and alterations in glucose homeostasis.

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Quantitative Proteomic Approaches in Mouse: Stable Isotope Incorporation by Metabolic (SILAC) or Chemical Labeling (Reductive Dimethylation) Combined with High‐Resolution Mass Spectrometry
Anja M. Billing, Hisham Ben Hamidane, Johannes Graumann

Mass spectrometry–based quantitative proteomics is a powerful method for in‐depth exploration of protein expression, allowing researchers to probe its regulation and study signal‐transduction networks, protein turnover, secretion, and spatial distribution, as well as post‐translational modi.

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Mouse Models of Bone Healing: Fracture, Marrow Ablation, and Distraction Osteogenesis
Kyle Lybrand, Beth Bragdon, Louis Gerstenfeld

Three commonly used murine surgical models of bone healing [closed fracture with intramedullary fixation, distraction osteogenesis (DO), and marrow ablation by reaming] are presented. Detailed surgical protocols for each model are outlined.

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Overview of Biological Mechanisms and Applications of Three Murine Models of Bone Repair: Closed Fracture with Intramedullary Fixation, Distraction Osteogenesis, and Marrow Ablation by Reaming
Beth Bragdon, Kyle Lybrand, Louis Gerstenfeld

Fractures are one of the most common large‐organ, traumatic injuries in humans, and osteoporosis‐related fractures are the fastest growing health care problem of aging.

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Mouse Anesthesia and Analgesia
Sean Adams, Cholawat Pacharinsak

Providing anesthesia and analgesia for mouse subjects is a common and critical practice in the laboratory setting.

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Fluorous‐Assisted Synthesis of (E)‐5‐[3‐Aminoallyl]‐Uridine‐5′‐O‐Triphosphate
Anilkumar R. Kore, Bo Yang, Balasubramanian Srinivasan

An efficient, reliable method for the chemical synthesis of (E)‐5‐[3‐aminoallyl]‐uridine‐5′‐O‐triphosphate (AA‐UTP), starting from 5‐iodouridine, is described.

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Microwave‐Assisted Phosphitylation of DNA and RNA Nucleosides and Their Analogs
Tim Efthymiou, Ramanarayanan Krishnamurthy

Microwave‐assisted chemical phosphitylation of novel nucleoside analogs containing a ribulose sugar unit was successful with yields ranging from 50% to 79% using 2‐cyanoethyl‐N,N‐diisopropyl chlorophosphoramidite as the phosphitylating reagent.

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Characterization of Thioether‐Linked Protein Adducts of DNA Using a Raney‐Ni‐Mediated Desulfurization Method and Liquid Chromatography‐Electrospray‐Tandem Mass Spectrometry
Goutam Chowdhury, F. Peter Guengerich

This unit contains a complete procedure for the detection and structural characterization of DNA protein crosslinks (DPCs). The procedure also describes an approach for the quantitation of the various structurally distinct DPCs.

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Synthesis of γ‐Phosphate‐Labeled and Doubly Labeled Adenosine Triphosphate Analogs
Stephan M. Hacker, Moritz Welter, Andreas Marx

This unit describes the synthesis of γ‐phosphate‐labeled and doubly labeled adenosine triphosphate (ATP) analogs and their characterization using the phosphodiesterase I from Crotalus adamanteus (snake venom phosphodiesterase; SVPD).

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Symmetrical Diamidate Prodrugs of Nucleotide Analogues for Drug Delivery
Fabrizio Pertusati, Christopher McGuigan, Michaela Serpi

The use of pronucleotides to circumvent the well‐known drawbacks of nucleotide analogs has played a significant role in the area of antiviral and anticancer drug delivery.

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Arrestin‐3‐Dependent Activation of c‐Jun N‐Terminal Kinases (JNKs)
Xuanzhi Zhan, Seunghyi Kook, Tamer S. Kaoud, Kevin N. Dalby, Eugenia V. Gurevich, Vsevolod V. Gurevich

Only one out of four mammalian arrestin subtypes, arrestin‐3, facilitates the activation of JNK family kinases. Here we describe two different protocols used for elucidating the mechanisms involved.

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Single‐Channel Recording of Glutamate Receptors
Chris Shelley

Highlighted in this unit are issues that should be considered when recording glutamate receptors at the single‐channel level, including some commonly encountered problems and their remedies. “UNIT 11.

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Single‐Channel Analysis of Glutamate Receptors
Chris Shelley

This is a companion to UNIT 11.16: Single‐Channel Recording of Glutamate Receptors. Described here are techniques for analyzing single‐channel currents recorded from glutamate receptors to characterize their properties.

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Generation of Human Acute Lymphoblastic Leukemia Xenografts for Use in Oncology Drug Discovery
Linda Holmfeldt, Charles G. Mullighan

The establishment of reproducible mouse models of acute lymphoblastic leukemia (ALL) is necessary to provide in vivo therapeutic test systems that recapitulate human ALL, and for amplification of limited amounts of primary tumor material.

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