1Department of Chemistry, University of Virginia
A suite of colorimetric assays is described for rapidly distinguishing protein, RNA, DNA, and reducing sugars in potentially heterogeneous biomolecular samples.
Published February 4, 2013. Keywords: Chemistry, Biochemistry, Chemical Biology, Genetics, Molecular Biology, Cellular Biology, Nucleic Acids, DNA, RNA, Proteins, analytical chemistry, Benedict's assay, Bial's orcinol assay, Dische's diphenylamine assay, colorimetric assay, reducing sugar, purification, transcription, reaction, assay
1Department of Chemical and Biological Engineering, Iowa State University, 2Department of Chemistry, Iowa State University
In this article, a high throughput method is presented for the synthesis of oligosaccharides and their attachment to the surface of polyanhydride nanoparticles for further use in targeting specific receptors on antigen presenting cells.
Published July 6, 2012. Keywords: Bioengineering, Chemical Engineering, High-throughput, Automation, Carbohydrates, Synthesis, Polyanhydrides, Nanoparticles, Functionalization, Targeting, Fluorous Solid Phase Extraction
1Great Lakes Bioenergy Research Center, Michigan State University (MSU), 2Great Lakes Bioenergy Research Center and DOE-Plant Research Lab, Michigan State University (MSU)
Plant biomass is a major carbon-neutral renewable resource that could be used for the production of biofuels. Plant biomass consists mainly of cell walls, a structurally complex composite material termed lignocellulosics. Here we describe a protocol for a comprehensive analysis of the content and composition of wall derived carbohydrates.
Published March 12, 2010. Keywords: Plant Biology, cell walls, polysaccharide, cellulose, hemicellulose, sugar composition, GC-MS
1Department of Bioengineering, Stanford University, 2Department of Molecular Biology and Laboratory for Molecular Infection Medicine Sweden, Umeå Centre for Microbial Research, Umeå University, 3Campus de Cantoblanco, Universidad Autonoma de Madrid, 4Department of Microbiology and Immunology, Stanford University School of Medicine
The bacterial cell wall is composed of peptidoglycan, a macromolecular network of sugar strands crosslinked by peptides. Ultra Performance Liquid Chromatography provides high resolution and throughput for novel discoveries of peptidoglycan composition. We present a procedure for the isolation of cell walls (sacculi) and their subsequent preparation for analysis via UPLC.
Published January 15, 2014. Keywords: Chemistry, peptidoglycan, bacterial cell wall, ultra-performance liquid chromatography, high-performance liquid chromatography, cell shape, morphogenesis
1Department of Cell Biology, University of Texas Southwestern Medical Center at Dallas
Procedures for complete reconstitution of a prototype voltage-gated potassium channel into lipid membranes are described. The reconstituted channels are suitable for biochemical assays, electrical recordings, ligand screening and electron crystallographic studies. These methods may have general applications to the structural and functional studies of other membrane proteins.
Published July 13, 2013. Keywords: Molecular Biology, Biochemistry, Genetics, Cellular Biology, Structural Biology, Biophysics, Membrane Lipids, Phospholipids, Carrier Proteins, Membrane Proteins, Micelles, Molecular Motor Proteins, life sciences, biochemistry, Amino Acids, Peptides, and Proteins, lipid-protein interaction, channel reconstitution, lipid-dependent gating, voltage-gated ion channel, conformation-specific ligands, lipids
1Natural Resource Ecology Laboratory, Colorado State University, 2Biosciences Division, Oak Ridge National Laboratory, 3Department of Bioengineering, University of Colorado
To measure potential rates of soil extracellular enzyme activities, synthetic substrates that are bound to a fluorescent dye are added to soil samples. Enzyme activity is measured as the fluorescent dye is released from the substrate by an enzyme-catalyzed reaction, where higher fluorescence indicates more substrate degradation.
Published November 15, 2013. Keywords: Environmental Sciences, Ecological and Environmental Phenomena, Environment, Biochemistry, Environmental Microbiology, Soil Microbiology, Ecology, Eukaryota, Archaea, Bacteria, Soil extracellular enzyme activities (EEAs), fluorometric enzyme assays, substrate degradation, 4-methylumbelliferone (MUB), 7-amino-4-methylcoumarin (MUC), enzyme temperature kinetics, soil
1Biochemistry and Molecular Biology, Michigan State Universtiy
Knowledge of the composition of the phloem sap as well as the mechanism of its loading and long-distance transport is essential for the understanding of long-distance signaling in plant development and stress/pathogen response and of assimilate transport. This manuscript describes the collection of phloem exudates using the EDTA-facilitated method.
Published October 23, 2013. Keywords: Plant Biology, plant, long-distance transport, long-distance signaling, phloem, phloem exudate collection, assimilate transport, protein, RNA, lipids
JoVE Immunology and Infection
1Undergraduate Program, Rice University, 2Proteomics Facility, Department of Pathology, University of Texas MD Anderson Cancer Center, 3Department of Melanoma Medical Oncology, University of Texas MD Anderson Cancer Center, 4University of Texas Graduate School of Biological Sciences at Houston
A specific and sensitive method to gain insight into the expression profile of glycosphingolipid antigens in immune organs and cells is described. The method takes advantage of the ion trap mass spectrometry allowing step-wise fragmentation of glycosphingolipid molecules for structural analysis in comparison to chemically synthesized standards.
Published April 16, 2013. Keywords: Immunology, Biochemistry, Molecular Biology, Cellular Biology, Structural Biology, Medicine, Genetics, Proteomics, Proteins, Glycomics, Functional glycomics, glycosphingolipids, GSLs, antigens, natural killer T cells, MALDI-TOF mass spectrometry, LTQ ion trap mass spectrometer, mass spectrometry, glycolipids, lipids
1BioSentinel Inc., Madison, WI
The BoTest Matrix botulinum neurotoxin (BoNT) detection assays rapidly purify and quantify BoNT from a range of sample matrices. Here, we present a protocol for the detection and quantification of BoNT from both solid and liquid matrices and demonstrate the assay with BOTOX, tomatoes, and milk.
Published March 3, 2014. Keywords: Neuroscience, Botulinum, food testing, detection, quantification, complex matrices, BoTest Matrix, Clostridium, potency testing
1Department of Biology, New Mexico State University
A ribosomal RNA (rRNA) depletion protocol was developed to enrich messenger RNA (mRNA) for RNA-seq of the mosquito gut metatranscriptome. Sample specific rRNA probes, which were used to remove rRNA via subtraction, were created from the mosquito and its gut microbes. Performance of the protocol can result in the removal of approximately 90-99% of rRNA.
Published April 7, 2013. Keywords: Genetics, Infection, Infectious Diseases, Molecular Biology, Cellular Biology, Microbiology, Genomics, biology (general), genetics (animal and plant), life sciences, Eukaryota, Bacteria, metagenomics, metatranscriptome, RNA-seq, rRNA depletion, mRNA enrichment, mosquito gut microbiome, RNA, DNA, sequencing