Rapid capillary zone electrophoresis in isoelectric histidine buffer: high resolution of the poly-T tract allelic variants in intron 8 of the CFTR gene

Clin Chem. 1998 May;44(5):906-13.

Abstract

The poly-T tract in intron 8 of the cystic fibrosis conductance transmembrane regulator (CFTR) gene exists in three variants, 5T, 7T, and 9T. The 7T and 9T variants generate a predominantly normal transcript, whereas the 5T variant engenders an anomalous product. The analysis of the poly-T tract is assuming increasing relevance, both to assess the implication of the CFTR gene in congenital bilateral absence of the vas deferens and to evaluate genotype-phenotype correlation in cystic fibrosis. Mapping of the poly-T tract has been performed by cumbersome and time-consuming methodologies. Capillary zone electrophoresis, combined with laser-induced fluorescence detection, was introduced for a rapid separation of the poly-T tract amplified products. As separation buffer, we adopted 200 mmol/L histidine (pH = pI = 7.6), and the capillary was filled with 10% polyacrylamide, allowing separations in less than 10 min. Capillary zone electrophoresis results were in perfect agreement with dot-blot analysis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles*
  • Buffers
  • Child
  • Cystic Fibrosis / genetics
  • Cystic Fibrosis / pathology
  • Cystic Fibrosis Transmembrane Conductance Regulator / genetics*
  • Electrophoresis, Capillary
  • Fluorescence
  • Histidine*
  • Humans
  • Introns / genetics*
  • Lasers
  • Male
  • Poly T / genetics*
  • Polymerase Chain Reaction
  • Vas Deferens / abnormalities

Substances

  • Buffers
  • CFTR protein, human
  • Cystic Fibrosis Transmembrane Conductance Regulator
  • Poly T
  • Histidine

Grants and funding