hybridized 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: Genetics and Genomics: Cancer Genetics Protocols

Anatomy of a comparative gene expression study. Includes: Choosing Cell Populations; mRNA Extraction and Reverse Transcription; Fluorescent Labeling of cDNA's; Hybridization to a DNA Microarray; Scanning the Hybridized Array; Interpreting the Scanned Image. - [Read Anatomy of a Comparative Gene Expression Study]

Category: Microarray Protocols: Microarray Analysis Protocols

Information on anatomy of a comparative gene expression study.  Includes: Choosing cell populations; mRNA Extraction and Reverse Transcription; Fluorescent labeling of cDNA's; Hybridization of DNA microarray; Scanning the hybridized array; Interpreting the scanned image. - [Read Anatomy of a Comparative Gene Expression Study]

Category: Cytogenetics Protocols: Fluorescent In Situ Hybridization FISH Protocol

Protocol for fluorescence in situ hybridization of a repetitive DNA probe to human chromosomes in suspension. Hybridization technique which does not need formamide and dextran sulfate. As a model system, we used the repetitive specific human DNA probe pUC 1.77, labeled it with digoxigenin-11-dUTP by nick-translation, and hybridized it to metaphase chromosomes in suspension. These chromosomes were isolated by standard techniques from human lymphocytes. - [Read Fluorescence In Situ Hybridization of a Repetitive DNA Probe to Human Chromosomes in Suspension]

Category: Nucleic Acid Purification Protocols

Preparations of RNA containing an mRNA of interest are hybridized to a complementary single-stranded DNA probe.  At the end of the reaction nuclease S1 is used to degrade unhybridized regions of the probe, and the surviving DNA-RNA hybrids are then separated by gel electrophoresis and visualized by autoradiography or Southern hybridization.  Method used to quantitate RNAs, to map the positions of introns, and to identify the locations of 5' and 3' ends of mRNAs on cloned DNA templates. - [Read Mapping RNA with Nuclease S1 Protocol]

Category: Nucleic Acid Modification Protocols: Nucleic Acid Radiolabeling Protocols

Preparations of RNA containing an mRNA of interest are hybridized to a radiolabeled single-stranded RNA probe. The method can be used to quantitate RNAs, to map the positions of introns, and to identify the locations of 5' and 3' ends of mRNAs on cloned DNA templates. - [Read Mapping RNA with Ribonuclease and Radiolabeled RNA Probes Protocol]