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Expression of Cloned Genes in E. coli Using the Bacteriophage lambda pL Promoter Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4086

Category: Viral Manipulation Protocols: Phage Lamda Manipulation Protocols

Protocol describes how (1) to clone cloned sequences encoding open reading frames in plasmids carrying bacteriophage {lambda} pL promoters, (2) to optimize expression of target proteins in transformants carrying these recombinants, and (3) to scale-up production of foreign proteins. - [Read Expression of Cloned Genes in E. coli Using the Bacteriophage lambda pL Promoter Protocol]

Expression of Cloned Genes in E. coli Using the Bacteriophage lambda pL Promoter Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot4086

Category: E Coli Protocols: E coli Protein Protocols

Protocol for expression of cloned genes in E. coli using the bacteriophage lambda pL promoter. Protocol describes how (1) to clone cloned sequences encoding open reading frames in plasmids carrying bacteriophage lambda pL promoters, (2) to optimize expression of target proteins in transformants carrying these recombinants, and (3) to scale-up production of foreign proteins. - [Read Expression of Cloned Genes in E. coli Using the Bacteriophage lambda pL Promoter Protocol]

Expression of Cloned Genes in E. coli Using the Bacteriophage T7 Promoter Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3597

Category: E Coli Protocols: E coli Protein Protocols

Protocol for the expression of cloned genes in E. coli using the bacteriophage T7 promoter. Protocol describes how (1) to clone cloned sequences encoding open reading frames in plasmids carrying bacteriophage T7 promoters, (2) to optimize expression of target proteins in transformants carrying these recombinants, and (3) to scale-up production of foreign proteins. - [Read Expression of Cloned Genes in E. coli Using the Bacteriophage T7 Promoter Protocol]

Expression Protocol in M9 Minimal Media via T7 Promoter - http://structbio.vanderbilt.edu/chazin/wisdom/labpro/M9.html

Category: Bacterial Cell Culture Protocols: Bacterial Cell Culture Media Protocols

This protocol has been used successfully to 15N or 13C/15N label our proteins using our pET1120/BL21(DE3) expression system: Preparing M9 minimal media begins with preparing a 5x stock solution of M9 salts. Generally, M9 salts contain a nitrogen source in the form of NH4Cl. Since we want to add a labeled nitrogen source, our 5x salts are prepared minus NH4Cl. Standard 5 X M9 Minimal Media salts minus nitrogen source For 1L 5xM9 salts: - [Read Expression Protocol in M9 Minimal Media via T7 Promoter]

Extraction and analysis of diagnostically useful proteins from formalin-fixed, paraffin-embedded tis - http://www.jhc.org/cgi/content/full/46/3/397

Category: Protein Protocols: Protein Extraction From Tissue

Extraction and analysis of diagnostically useful proteins from formalin-fixed, paraffin-embedded tissue sections. Ikedaa et al, 1998 JHC. - [Read Extraction and analysis of diagnostically useful proteins from formalin-fixed, paraffin-embedded tis]

Extraction of Bacteriophage lambda DNA from Large-scale Cultures Using Formamide Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3973

Category: Viral Manipulation Protocols: Phage Lamda Manipulation Protocols

Formamide can be used instead of proteinase K to dissociate bacteriophage {lambda} coat proteins from the viral {lambda}. The procedure is rapid and works best with large-scale preparations of bacteriophage {lambda}. - [Read Extraction of Bacteriophage lambda DNA from Large-scale Cultures Using Formamide Protocol]

Extraction of Bacteriophage lambda DNA from Large-scale Cultures Using Proteinase K and SDS Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3972

Category: Viral Manipulation Protocols: Phage Lamda Manipulation Protocols

DNA is best isolated from bacteriophage lambda particles by digesting the viral coat proteins with a powerful protease such as proteinase K, followed by extraction with phenol:chloroform. This procedure is easily adapted for use with smaller-scale preparations of bacteriophage lambda. - [Read Extraction of Bacteriophage lambda DNA from Large-scale Cultures Using Proteinase K and SDS Protocol]

Extraction of Total Protein from Arabidopsis Protocol - http://www.cshprotocols.org/cgi/content/abstract/2007/1/pdb.prot4680

Category: Plant Biology Protocols: Arabidopsis Protein Peptide Protocols

The simplest way to analyze proteins is in unfractionated extracts. However, it is often desirable to fractionate proteins, e.g, by size. This procedure extracts total protein from Arabidopsis samples. Typical yields are ~2-3 mg/ml (using rosette leaves) or 6-8 mg/ml (using young seedlings). - [Read Extraction of Total Protein from Arabidopsis Protocol]

Fixing Attached Cells in Organic Solvents Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/21/pdb.prot4293

Category: Cell Culture Protocols: Cell Fixing Protocols

Organic solvents such as alcohols and acetone remove lipids and dehydrate cells, precipitating the proteins on the cellular architecture. Be aware that different antigens may be affected differently by the various solvents. If no previous data are available for your antigen, start with the 50/50 mixture. For tissue culture dishes, concentrations of acetone higher than 50% will destroy the integrity of the plastic. - [Read Fixing Attached Cells in Organic Solvents Protocol]

Fluorescein Labeling of Proteins Protocol - http://pingu.salk.edu/flow/protocols/flabel.html

Category: Cell Biology and Cell Culture: Flow Cytometry Protocols: Flow Cytometry Immunofluorescence Staining Protocols

Protocol for fluorescein labeling of proteins. - [Read Fluorescein Labeling of Proteins Protocol]

Fractional Precipitation of Proteins by Ammonium Sulfate - Subscription - http://www.cshprotocols.org/cgi/content/full/2006/16/pdb.prot4309

Category: Protein Protocols: Protein Precipitation Procols

Fractional Precipitation of Proteins by Ammonium Sulfate Simpson Cold Spring Harbor Protocols.2006; 2006: pdb.prot4309 - [Read Fractional Precipitation of Proteins by Ammonium Sulfate - Subscription]

Fractionation of Maize Embryo Proteins for 2-D Gel Electrophoresis Using Multicompartment Electrolyz - http://www.cshprotocols.org/cgi/content/extract/2006/1/pdb.prot4235

Category: Plant Biology Protocols: Plant Proteins, Peptides and Antigens

Protocol describes a method for performing isoelectric fractionation of a maize embryo sample using a multicompartment electrolyzer(MCE). This prefractionation of proteins having pIs within a certain pH interval is essential for allowing high loads of protein to be resolved on narrow and ultra-narrow immobilized pH gradients used in 2D electrophoresis. The isoelectric membranes in the MCE act like isoelectric traps capturing all the protein species having pIs encompassing the pI value of each... - [Read Fractionation of Maize Embryo Proteins for 2-D Gel Electrophoresis Using Multicompartment Electrolyz]

Gel Retardation Assays for DNA-binding Proteins Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3948

Category: Nucleic Acid Electrophoresis Protocols: DNA Electrophoresis Protocols: DNA Electrophoresis in Polyacrylamide Gels Protocols

Protocol exploits differences in electrophoretic mobility through a nondenaturing polyacrylamide gel between a rapidly migrating target DNA and a more slowly migrating DNA-protein complex. - [Read Gel Retardation Assays for DNA-binding Proteins Protocol]

Gene Expression and Protein Localization in Arabidopsis Protocols - http://www.arabidopsis.org/cshl-course/7-gene_expression.html

Category: Plant Biology Protocols: Arabidopsis Genetics Protocols

Protocols for gene expression and protein localization in Arabidopsis. Includes: Detection of the native protein; Detection of a recombinant version; Immunofluorescence detection in Arabidopsis protoplasts; Isolation of Arabidopsis seedling protoplasts; Subcellular localization of GUS-fusion proteins in Arabidopsis seedlings; Localization of Arabidopsis proteins with GUS in situ enzyme assay. - [Read Gene Expression and Protein Localization in Arabidopsis Protocols]

General Overview of Histology - http://www-bioc.rice.edu/bios576/histology/histology.htm

Category: Histology Protocols: Fixation of Cells Tissues Protocol

Fixation must convert the soluble contents of the cell (ie proteins, carbohydrates, ... http://www.protocol-online.org/prot/Histology/Fixation/ ...General Overview of Histology and Tissue Fixation. BIOS 576. Rice. - [Read General Overview of Histology]

Gentle lysis of mammalian cells for cytochrome c release assay - http://www.celldeath.de/apometh/mitobuff.html

Category: Cell Biology and Cell Culture: Apoptosis Protocols

Lysis in the sucrose-containing Buffer ("Mito-Buffer") is supposed to prevent accidential disrupture of the mitochondria to prevent the leakage of mitochondrial proteins (such as cytochrome c) into the cytosol. Celldeath.de - [Read Gentle lysis of mammalian cells for cytochrome c release assay]

Gold labeling of proteins for electron microscopy - http://www.biochem.wisc.edu/inman/empics/gold.htm

Category: Molecular Imaging: Electron Microscopy Protocols

In brief, the reaction mixture, is prepared in exactly the same way as if the sample is to be stained or shadowed for electron microscopy, but once the... Gold labeling technique for electron microscopic identification and location of proteins. - [Read Gold labeling of proteins for electron microscopy]

Hamster Ovary Cell NA+/K+ -ATPase Test - http://embryo.ib.amwaw.edu.pl/invittox/prot/35.htm

Category: Pharmacology and Toxicology Protocols: Cytotoxicity Protocols: Cell Specific Cytotoxicity Assays Protocols

Accumulation of lipophilic substances in the plasma membrane may affect the membrane lipid order and consequently affect the function of these proteins. Changes in the activity of the Na+/K+ -ATPase, which is the major active transport system responsible for the electrochemical potential in mammalian cells, can therefore be an indication of the effect that a chemical may have on the viability of the cell membrane and possibly the whole cell. - [Read Hamster Ovary Cell NA+/K+ -ATPase Test]

His Tag Nickel Affinity Chromatography Protocol PDF - http://www.mnstate.edu/provost/His_TagNiChromatogProtocol.pdf

Category: Chromatography Protocols

His Tag Nickel Affinity Chromatography Protocol PDF. The Wallert and Provost Lab. Theory and Introduction: Ni-Affinity Chromatography uses the ability of His to bind nickel. Six histadine amino acids at the end of a protein (either N or C terminus) is known as a 6X His tag. Nickel is bound to an agarose bead by chelation using nitroloacetic acid (NTA) beads. Several companies produce these beads as His Tagged proteins are some of the most used affinity tags in todayâ€™s market. - [Read His Tag Nickel Affinity Chromatography Protocol PDF]

How to Cross-Link Proteins - http://www.fgsc.net/neurosporaprotocols/How%20to%20cross-link%20proteins.pdf

Category: Protein Protocols: Immunoprecipitation Protocols

Information and tips on how to cross-link proteins. - [Read How to Cross-Link Proteins]

How to Immunoblot Proteins - http://www.fgsc.net/neurosporaprotocols/how%20to%20immunoblot%20proteins.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to immunoblot proteins. - [Read How to Immunoblot Proteins]

How to Isolate Proteins - http://www.fgsc.net/neurosporaprotocols/How%20to%20isolate%20proteins%20final.pdf

Category: Fungi Protocols: Neurospora Protocols

Information and tips on how to isolate proteins. - [Read How to Isolate Proteins]

How to Use GFP-Tagged Proteins in Cell Biology - http://www.fgsc.net/neurosporaprotocols/How%20to%20use%20GFP-tagged%20proteins%20in%20cell%20biology.pdf

Category: Reporter Gene Assays: GFP Assay Protocols

Information and tips on how to use GFP-tagged proteins in cell biology - [Read How to Use GFP-Tagged Proteins in Cell Biology]

HPLC Separation of Digested Proteins and Preparation Protocol - http://cshprotocols.cshlp.org/cgi/content/abstract/2006/30/pdb.prot4663

Category: Chromatography Protocols: Liquid Chromatography Protocols

Protocol for HPLC separation of digested proteins and preparation for matrix-assisted laser desorption/ionization analysis. - [Read HPLC Separation of Digested Proteins and Preparation Protocol]

Identification of Associated Proteins by Coimmunoprecipitation Protocol - http://www.cshprotocols.org/cgi/content/extract/2006/2/pdb.prot3898

Category: Immunology: Immunoprecipitation Protocols: Immunoprecipitation of Radiolabeled Cells Protocols

Coimmunoprecipitation is most commonly used to test whether two proteins of interest are associated in vivo, but it can also be used to identify novel interacting partners of a target protein. In both cases, the cells, which may have been labeled with [35S]methionine, are harvested and lysed under conditions that preserve protein-protein interactions. The target protein is specifically immunoprecipitated from the cell extracts, and the immunoprecipitates are fractionated by SDS-PAGE. - [Read Identification of Associated Proteins by Coimmunoprecipitation Protocol]

Articles (1-3 of 3)

Acid Guanidinium Thiocyanate RNA Isolation Phenol Chloroform Extraction

A single step RNA isolation protocol using Phenol Chloroform Extraction and Acid Guanidinium Thiocyanate. This RNA isolation method uses the fact that guanidinium thiocyanate can simultaneously lyse the cells and inactive cellular RNAses during the initial RNA isolation step allow a single step in the method.

In Vitro Translated Xenopus Mos Kinase Assay Protocol

In Vitro Translated Xenopus Mos Kinase Assay Protocol. In response to progesterone, immature Xenopus oocytes mature to eggs that can be fertilized. The Mos protein kinase is essential for oocyte maturation, most likely due to its ability to activate the MAP kinase cascade. This MAP kinase cascade eventually leads to the activation of Cdc2/cyclin B and entry into M phase. In this protocol, tagged Mos kinase is translated in vitro, immunopurified, and used in a kinase assay.