structure Protocols

Category: PCR Protocols: Reverse Transcriptase RT-PCR Protocols

Protocol uses a single thermostable RNA polymerase to perform high-specificity RT-PCR.  A high-temperature RT reaction is followed by PCR amplification of the cDNA using a single thermostable poymerase, the GeneAmp AccRT RNA PCR enzyme from Applied Biosystems.  The high temperature of the RT reaction enhances the specificity of primer binding and also reduces secondary structure in the template, thereby increasing the efficiency of polymerization. - [Read Amplification of RNA: High-Temperature Reverse Transcription and DNA Amplification with a Magnesium]

Category: Cell Culture Protocols: Sterile Technique Protocols

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. This unit describes the basic skills required to maintain and preserve cell cultures: aseptic technique, medium characteristics, passaging, freezing and storage, recovering frozen stocks, and counting viable cells. - [Read Basic Techniques for Mammalian Cell Tissue Culture Protocol]

Category: Primary Cell Isolation and Culture Protocols: Mammalian Cell Culture Protocols

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. This unit describes the basic skills required to maintain and preserve cell cultures: aseptic technique, medium characteristics, passaging, freezing and storage, recovering frozen stocks, and counting viable cells. - [Read Basic Techniques for Mammalian Cell Tissue Culture Protocol]

Category: PCR Protocols: cDNA Synthesis Protocols

This method, for the selective amplification of full-length cDNA ends, involves the addition of an adapter during reverse transcription.  This method takes advantage of the propensity of Moloney murine leukemia virus reverse transcriptase (MMLV RT) to append two to four cytosines to the 3'-end of newly synthesized cDNA strands.  The additional residues are added when the enzyme reaches the 5'-cap structure at the end of the mRNA template. - [Read Cap-Switching RACE Protocol]

Category: DNA Protocols: DNA Sequencing Protocols: Dideoxy Mediated Sequencing Protocols

Because sequencing reactions catalyzed by thermostable DNA polymerases such as Taq are carried out at elevated temperatures, problems caused by mismatched annealing of primers or templates rich in secondary structure are greatly alleviated. - [Read Dideoxy-mediated Sequencing of DNA Using Taq DNA Polymerase Protocol]

Category: Immunology: Immunofluorescence Protocols: Immunofluorescence Staining Protocols

Provides a protocol for indirect immunofluorescence, which is a method that provides information about the locations of specific molecules and the structure of the cell. Antibody molecules for a specific target molecule are exposed to the cell or tissue being investigated. The binding of these molecules is detected by incubating the sample with a secondary antibody specific for immunoglobulin molecules and conjugated to fluorophore. - [Read Immunofluorescence Staining Protocol]

Category: Primary Cell Isolation and Culture Protocols: Laser Capture Microdissection Protocols

Useful techniques to circumvent disruption of tissue structure in the analysis of gene expression are LCM and LDM. While they require specialized microscopes and systems, they are similar in that freshly-cut frozen tissue sections can be microdissected using either a general histological stain (like H&E) or by staining with fluorescently conjugated antibodies. The LCM system by Arcturus involves... - [Read Immunofluorescent Staining for the Laser Microdissection of Individual Cells Protocol]

Category: Nucleic Acid Modification Protocols: DNA Methylation Protocols

Methods for analyzing the role of DNA methylation and chromatin structure in regulating T lymphocyte gene expression. - [Read Methods for Analyzing the Role of DNA Methylation and Chromatin Structure in Regulating T Lymphocyte]

Category: Neuroscience Protocols

Methods for structure function studies of neurotrophic factors. - [Read Methods for Structure Function Studies of Neurotrophic Factors]

Category: Cytoskeleton Protocols: Microtubules Protocols

Negative staining is a rapid, qualitative method for analyzing microtubule structure at the EM level. Because negative staining involves deposition of heavy atom stains, structural artifacts such as flattening of the cylindrical microtubule and opening up of microtubules into flat sheets are common. Negative staining is very useful because of its ease, rapidity and lack of requirement for specialized equipment other than that found in a regular EM facility. - [Read Negative Stain Electron Microscopy of Microtubules Protocol]

Category: Neuroscience Protocols

This tutorial has images in which the structures are labelled.  You are to identify the structures by clicking on the name of the structure. - [Read Neuroanatomy Tutorial]

Category: DNA Protocols: DNA Sequencing Protocols

The inclusion of formamide in sequencing gels eliminates secondary structure in the DNA during electrophoresis. Formamide gels are particularly useful and almost a necessity when sequencing DNA templates with a G/C content >55%. - [Read Preparation of Denaturing Polyacrylamide Gels Containing Formamide Protocol]

Category: Nucleic Acid Electrophoresis Protocols

Protocol for preparation of denaturing polyacrylamide gels containing formamide.The inclusion of formamide in sequencing gels eliminates secondary structure in the DNA during electrophoresis.  Formamide gels are particularly useful and almost a necessity when sequencing DNA templates with a G/C content >55%. - [Read Preparation of Denaturing Polyacrylamide Gels Containing Formamide Protocol]

Category: Immunohistochemistry Protocols: Antibody Staining Protocols

Frozen tissue sections show good preservation of tissue structure and antigens. The principle disadvantages of using them in immunostaining are that the specimens must be stored frozen, and a special microtome, known as a cryostat, is required. Also, many clinical specimens are not available in this form, and most classic histological descriptions of tissue structure and pathology are based on the use of paraffin-embedded sections of formalin-fixed material. - [Read Preparing Frozen Tissue Sections for Immunostaining Protocol]