title: Quantification of the mRNA expression of G protein-coupled receptors in human adipose tissue.
abstract: G protein-coupled receptors (GPCRs) are important regulators of human physiology and therefore the targets of a large number of modern therapeutics. Although GPCRs are important regulators of adipose tissue endocrine and energy storage functions, the expression and function of a majority of GPCRs in adipose tissue is poorly characterized. A first step in the functional characterization of adipose tissue GPCRs is to accurately quantify the expression of GPCRs in adipose tissue. In this methods chapter, a detailed, step-by-step protocol is presented for the isolation of adipose tissue total RNA, its conversion into cDNA and the real-time PCR quantification of human GPCR mRNA expression relative to the mRNA expression of the stable adipose tissue housekeeping gene peptidylprolyl isomerase A (PPIA). A comprehensive list of 377 manually validated, commercially available GPCR qPCR primers allows facilitated swift quantification of either the entire human GPCRome or individual GPCRs, thus providing a sensitive, flexible, and cost-effective means of determining the mRNA expression of GPCRs in adipose tissue.
authors.affiliation: Diabetes Research Group, King's College London, London, UK. Electronic address: stefan.amisten@kcl.ac.uk.
authors.forename: Stefan
authors.initials: S
authors.lastname: Amisten
mesh_terms: D020869:Gene Expression Profiling; D006801:Humans; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D060888:Real-Time Polymerase Chain Reaction; D043562:Receptors, G-Protein-Coupled / Q000235:genetics / Q000378:metabolism*; D050151:Subcutaneous Fat / Q000378:metabolism*; D059467:Transcriptome
chemical_list: D012333:RNA, Messenger; D043562:Receptors, G-Protein-Coupled
keywords: ADRA2A; Adipose; Drug targets; G protein-coupled receptors; GPCR; GPCRome; Microarray; PPIA; Quantitative real-time PCR; RNA-sequencing; qPCR