essentially Protocols

Category: PCR Protocols: cDNA Synthesis Protocols

An oligodeoxynucleotide primer hybridized to mRNA is extended by an RNA-dependent DNA polymerase to create a cDNA copy that can be amplified by PCR.  Depending on the purpose of the experiment, the primer for first-strand cDNA synthesis can be specifically designed to hybridize to a particular target gene, or a general primer such as oligo(dT) can be used to prime cDNA synthesis from essentially all mammalian mRNAs - [Read Amplification of cDNA Generated by Reverse Transcription of mRNA Protocol]

Category: Viral Culture and Isolation Protocols: Viral Culture Protocols

There are essentially three parts to this protocol: 1. growth of at least 5x10e8 pfu phage to provide an inoculum growth of a larger liquid lysate that will produce about 5x10e12 pfu; 2. concentration and purification of the phage, and; 3. DNA preparation. - [Read Growth and Purification of 25-100 ug Lambda Clone DNA Protocol]

Category: Antibody Protocols: Antibody Labeling Protocols

This method for tagging monoclonal antibodies involves growing hybridomas in the presence of radioactive amino acids. This protocol can be particularly useful when conventional labeling techniques cause the antibody to lose activity. The labeled antibodies that result are essentially identical to the unlabeled antibodies. - [Read Labeling Monoclonal Antibodies by Biosynthesis Protocol]

Category: Protein Protocols: Protein Extraction From Tissue

Plant Protein Isolation - Rapid isolation of protein for SDS-PAGE analysis. (Essentially the same protocol as that described for GUS Assays). Stockinger Lab - [Read Plant Protein Isolation]

Category: Viral Manipulation Protocols: Phage Protocols

Protocol describes a method for generating isolated plaques from a stock of bacteriophage lambda. Each plaque derives from infection of a single bacterium by a single bacteriophage particle. Because each plaque contains the progeny of a single virus particle, the bacteriophages derived from a single plaque are essentially genetically identical to one another. - [Read Plating Bacteriophage Lambda Protocol]

Category: Viral Protocols

Isopycnic centrifugation through CsCl gradients is used to prepare infectious bacteriophage {lambda} particles of the highest purity that are essentially free of contaminating bacterial nucleic acids. - [Read Purification of Bacteriophage lambda Particles by Isopycnic Centrifugation through CsCl Gradients]

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols: Membrane Isolation Protocols

The first part of the isolation procedure is a flotation through a continuous iodixanol gradient; this gradient is essentially a resolving gradient in which the caveolin-rich vesicles are concentrated in the top third of the gradient, while the predominantly caveolin-poor vesicles band in denser regions. A second discontinuous gradient is essentially a concentration gradient to band the caveolin-rich vesicles sharply at an interface. - [Read Purification of Caveolae Membranes from a Plasma Membrane Fraction of Cultured Cells and Tissues]

Category: Cell Organelle Protocols: Subcellular Fractionation Protocols

In the routine method described in this protocol, chylomicron-free plasma is adjusted to 12% (w/v) iodixanol and the sample, essentially fills an approx 3 ml tube for a near-vertical rotor. During the centrifugation VLDL, LDL and HDL particles and also plasma proteins migrate from all parts of the sample to their final buoyant density banding position in the self-forming density gradient. - [Read Subfractionation of Low-Density Lipoprotein (LDL)]