Streamlined Approach to Functional Analysis of Promoter-Region Polymorphisms. (August 2002)

• Record Type: Journal Article
• Authors: Coleman, S.L.; Hoogendoorn, B.; Guy, C.; Smith, S.K.; O'Donovan, M.C.; Buckland, P.R.
• Is Part Of: Biotechniques. Volume 33:Number 2(2002)
• Page Start: 412
• Page End: 418
• Publication Date: 2002-08
• Subjects: Biology, Experimental -- Periodicals; Molecular biology -- Periodicals; Medical technology -- Periodicals; Biology, Experimental; Medical technology; Molecular biology; Clinical Laboratory Techniques -- Periodicals; Research -- Periodicals; Medical Laboratory Science -- Periodicals; Periodicals; Electronic journals; 570
• DOI: 10.2144/02332ht01
• Languages: English
• ISSNs: 0736-6205

Abstract

We have developed a rapid method for identifying functional promoter-region polymorphisms. Using a modified pGL3 luciferase expression T-vector, we can amplify by PCR, clone, identify allelic pairs of a polymorphic gene promoter region, and prepare plasmids for cell culture 10 promoters (20 allele pairs) per week per researcher. By utilizing 96-well plate technology and an internal control plasmid expressing secreted alkaline phosphatase, each of these allele pairs can be tested for relative promoter activity in each of three cell lines (HEK293t, TE671, and JEG3) with similar resources.