High resolution melting (HRM) analysis for the detection of ER22/23EK, BclI, and N363S polymorphisms of the glucocorticoid receptor gene

Paolo Maltese, Emanuele Canestrari, Linda Palma, Annamaria Ruzzo, Francesco Corini, Michele Menotta, Francesca Andreoni, Anna Latiano, Vito Annese, Mauro Magnani

Research output: Contribution to journalArticle

Abstract

Polymorphisms in the glucocorticoid receptor (GR) gene have been associated with altered sensitivity to glucocorticoids. We designed a high-resolution melting (HRM) assay to detect, simultaneously, the three most intriguing GR polymorphisms, selected on the bases of clinical relevance and frequencies in caucasian population as described in literature. HRM enables the detection of ER22/23EK and N363S genotypes but fails to discriminate homozygous mutant for the BclI polymorphism from wild-type samples, however a simple spike experiment leads to a clear discrimination between these genotypes. The analyses were performed on a cohort of 70 healthy Caucasian subjects. The method was validated by restriction fragment length polymorphisms; HRM results were found to be in 100% concordance with those observed with the restriction enzymes. We also employed this method on a population of 40 Crohn Disease patients; the analysis demonstrated a significantly higher frequency of the BclI polymorphism in patients than in healthy volunteers. This is, at now, the less expensive and time-and work-saving method to detect GR mutations, providing precision, fast screening and high throughput capabilities.

Original languageEnglish
Pages (from-to)269-274
Number of pages6
JournalJournal of Steroid Biochemistry and Molecular Biology
Volume113
Issue number3-5
DOIs
Publication statusPublished - Feb 2009

Keywords

  • BclI
  • Crohn Disease
  • ER22/23EK
  • Glucocorticoid receptor
  • High-resolution melt
  • N363S
  • Single nucleotide polymorphism

ASJC Scopus subject areas

  • Molecular Medicine
  • Endocrinology, Diabetes and Metabolism
  • Molecular Biology
  • Cell Biology
  • Endocrinology
  • Clinical Biochemistry
  • Biochemistry

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