[Abstract]  [Full Text]  [PDF]

Jack M. Gallup^1* and Mark R. Ackermann¹

¹ Department of Veterinary Pathology, College of Veterinary Medicine, Iowa State University. Ames, Iowa 50011-1250. USA.

* To whom correspondence should be addressed: Jack M. Gallup, Department of Veterinary Pathology, College of Veterinary Medicine, Iowa State University. Ames, Iowa 50011-1250. USA. Phone: 1-515-294-5423. Fax: 1-515-294-5844. Email: eag@iastate.edu

Biol. Proced. Online 2006;8:87-152. doi:10.1251/bpo122
Submitted: May 02, 2006; Accepted: July 10, 2006; Published: September 15, 2006.

Indexing terms: Reverse Transcription; DNA, Complementary.

The purpose of this manuscript is to discuss fluorogenic real-time quantitative polymerase chain reaction (qPCR) inhibition and to introduce/define a novel Microsoft Excel-based file system which provides a way to detect and avoid inhibition, and enables investigators to consistently design dynamically-sound, truly LOG-linear qPCR reactions very quickly. The qPCR problems this invention solves are universal to all qPCR reactions, and it performs all necessary qPCR set-up calculations in about 52 seconds (using a pentium 4 processor) for up to seven qPCR targets and seventy-two samples at a time – calculations that commonly take capable investigators days to finish. We have named this custom Excel-based file system "FocusField2-6GallupqPCRSet-upTool-001" (FF2-6-001 qPCR set-up tool), and are in the process of transforming it into professional qPCR set-up software to be made available in 2007. The current prototype is already fully functional.