A Screening System for the Human Adrenergic Receptor

As a consequence to the study outlined in the previous chapter, it was necessary to establish a fast method for detecting the described mutations of the 2ar. The receptor gene has proved to show an increased polymorphism , as the sequence analysis of only nine subjects revealed four different alleles. We hoped to be able to associate one or more of the found point mutations (see section results) with an established phenotype. Therefore, in an extension of the previous study, a scanning method for the detection of these mutations was developed, and the mutation pattern of persons with known receptor density was determined.

• Methods of Detection of Single Point Mutations
• Detection of Point Mutations by RFLP of PCR Amplified DNA Sequences
  • Restriction Map of the Adrenergic Receptor Codon
  • Amplified Fragments
  • Selecting the Used Restriction Endonucleases
• PCR of the RFPL fragments
  • Used Protocol
  • PCR Optimization
• RFLP Protocol
• Analysis of RFLPs
• Silver Staining of DNA in Polyacrylamide Gels
  • Silver Staining Protocol
• Results
  • Human Adrenergic Receptor Allele Frequencies and Haplotypes
  • Adrenoceptor Alleles in Relation to Adrenoceptor Expression in Cultured Skin Fibroblasts