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Rapid methods for population-scale analysis for gene polymorphisms: the ACE gene as an example.
  1. S. D. O'Dell,
  2. S. E. Humphries,
  3. I. N. Day
  1. Department of Medicine, University College London Medical School, Rayne Institute.

    Abstract

    OBJECTIVE--To obtain rapid, high throughput genotyping of the angiotensin converting enzyme (ACE) gene intron 16 insertion/deletion polymorphism. METHODS--DNA was obtained from whole blood samples by a simple liquid phase methanol extraction procedure. The ACE gene was amplified by the polymerase chain reaction (PCR) using two oligonucleotide primers (ACE1 and ACE3) outside the insertion sequence and one primer (ACE2) inside the sequence. Microtitre array diagonal gel electrophoresis (MADGE) was used to determine genotypes. RESULTS--84 and 65 bp PCR products indicating the presence of deletion (D) and insertion (I) alleles, respectively, were clearly resolved after electrophoresis on a 7.5% polyacrylamide gel. Up to 480 DNA samples on 5 gels could be genotyped in a single electrophoresis run, or up to 1000 samples in a working day. CONCLUSIONS--A simplified DNA extraction protocol coupled to the high throughput capability of the MADGE electrophoretic system for genotyping enables analysis of large populations for association studies of ACE genotype with cardiac disease events.

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