title: Radiolabeled semi-quantitative RT-PCR assay for the analysis of alternative splicing of interleukin genes.
abstract: Alternative splicing evolved as a very efficient way to generate proteome diversity from a limited number of genes, while at the same time modulating posttranscriptional events of gene expression-such as stability, turnover, subcellular localization, binding properties, and general activity of both mRNAs and proteins. Since the vast majority of human genes undergo alternative splicing, it comes to no surprise that interleukin genes also show extensive alternative splicing. In fact, there is a growing body of evidence indicating that alternative splicing plays a central role in modulating the pleiotropic functions of cytokines, and aberrant expression of alternatively spliced interleukin mRNAs has been linked to disease. However, while several interleukin splice variants have been described, their function is still poorly understood. This is particularly relevant, since alternatively spliced cytokine isoforms can act both as disease biomarkers and as candidate entry points for therapeutic intervention. In this chapter we describe a protocol that uses radiolabeled semi-quantitative RT-PCR to efficiently detect, analyze, and quantify alternative splicing patterns of cytokine genes.
authors.affiliation: Department of Biological Sciences, St. John's University, 8000 Utopia Parkway, Queens, NY, 11439, USA.
authors.forename: Felitsiya
authors.initials: F
authors.lastname: Shakola
authors.forename: Stephen
authors.initials: S
authors.lastname: Byrne
authors.forename: Kainaat
authors.initials: K
authors.lastname: Javed
authors.forename: Matteo
authors.initials: M
authors.lastname: Ruggiu
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D017398:Alternative Splicing; D002460:Cell Line; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D015851:Interleukin-7 / Q000235:genetics* / Q000378:metabolism; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000378:metabolism*; D058953:Neural Stem Cells / Q000166:cytology / Q000378:metabolism*; D010761:Phosphorus Radioisotopes; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism; D022821:RNA Splice Sites; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods*
chemical_list: D017931:DNA Primers; C508605:IL7 protein, human; D015851:Interleukin-7; D010761:Phosphorus Radioisotopes; D020033:Protein Isoforms; D022821:RNA Splice Sites; D000255:Adenosine Triphosphate