purposes Protocols

Category: Plant Biology Protocols: Plant Genetics Protocols

Microsatellite markers, also referred to as STMS (SequenceTagged Microsatellite Sites) or STR (Short Tandem Repeats) are widely used as molecular markers for intraspecific genotyping, molecular mapping and breeding purposes. The method described is an efficient,fast and relatively inexpensive way to obtain microsatellite markers without post-cloning selection methods. So far, the method has been successful in onion (Allium cepa L.), a plant with a large genome and for pathogenic fungi. - [Read Enrichment for Microsatellite Sequences in Onion (Allium cepa L.) Protocol]

Category: Viral Manipulation Protocols: M13 Phage Manipulation Protocols

Protocol used chiefly to generate large stocks of double-stranded DNA of strains of M13 that are routinely used as cloning vectors. Large amounts of single-stranded DNA of an individual recombinant may occasionally be needed for specific purposes, e.g., to generate many preparations of a particular radiolabeled probe or to construct large numbers of site-directed mutants. - [Read Large-scale Preparation of Single-stranded and Double-stranded Bacteriophage M13 DNA Protocol]

Category: Cell Biology and Cell Culture: Apoptosis Protocols: Apoptosis Analysis

Measurement of Apoptosis and Other Forms of Cell Death. Jagan Muppidi, Melissa Porter, and Richard M. Siegel. As programmed cell death (PCD) or apoptosis has emerged as an important regulator of development and homeostasis in multicellular organisms, methods to quantify apoptosis and to distinguish it from necrosis have been developed. This unit presents a set of assays for these purposes, many of which are technically very simple and ideally suited to the study of hematopoietic cells. - [Read Measurement of Apoptosis and Other Forms of Cell Death]

Category: Developmental Biology: Utilizing Model Organisms Protocols

Nonviral, DNA-mediated gene transfer is an alternative to viral delivery systems for expressing new genes in cells and tissues. The Sleeping Beauty (SB) transposon system combines the advantages of viruses and naked DNA molecules for gene therapy purposes; however, efficacious delivery of DNA molecules to animal tissues can still be problematic. Here we describe the hydrodynamic delivery procedure for the SB transposon system that allows efficient delivery to the liver in the mouse. - [Read Preferential Delivery of the Sleeping Beauty transposon System to Livers of Mice by Hydrodynamic i]

Category: Immunology: Mononuclear Cell Isolation Protocols: T-Cell Isolation Protocols

Protocols utilize T hybridomas to detect expression of peptide-MHC complexes, since these cells provide the most convenient, consistent, and flexible T cell readout systems for these purposes. If desired, antigen-specific T cell clones can be used in lieu of T hybridoma cells, but T cell clones often give poorer responses than T hybridomas to fixed APCs due to fixation-induced loss of costimulator function. - [Read Presenting Exogenous Antigen to T Cells Protocol]

Category: Protein Protocols: Protein Extraction From Tissue

Combination of nucleic acid and protein isolation with tissue array construction: Using defined histologic regions in single frozen tissue blocks for multiple research purposes - [Read Protein isolation with tissue array construction]

Category: Nucleic Acid Purification Protocols: RNA Isolation Protocols: RNA Isolation fromTissues or Cultured Cells Protocols

Protocol is the second in a set of three, describing fluorescent mRNA differential display (FDD or FDDRT-PCR).  For the purposes of FDD gene expression analysis, as well as any other RNA-based gene expression technologies, contaminating genomic DNA must be removed before reverse transcription and subsequent PCR. - [Read Removal of Genomic DNA from Total RNA for Use in Fluorescent mRNA Differential Display Protocol]

Category: Plant Biology Protocols

Alfalfa is an outcrossing species, cultivars consist of populations rather than individual homozygous or inbred lines. After only two cycles of self-pollination, severe inbreeding depression eliminates selfed individuals from populations. To obtain sufficient plant material of relative genetically uniformity (these plants will still necessarily be heterozygous) for experimental purposes, it may be necessary to propagate a single individual or clone. - [Read Vegetative Propagation of Alfalfa by Stem Cuttings]