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Comparison of Fluorometric and Spectrophotometric DNA Quantification for Real-time Quantitative PCR of Degraded DNA

Published

Author(s)

Luke A. Shokere, Marcia J. Holden, G. Ronald Jenkins

Abstract

Isogenic NK603 DNA was degraded by sonication or heat and quantified using A260 and a fluorescent dye method. qPCR experiments were conducted by amplifying an SSIIb-3 endogenous control and an NK603 transgene in untreated, sonicated, and heat-treated samples. qPCR reactions on sonicated DNA samples, based on A260 quantification, provided 0.125%, 1.14% and 2.15% NK603; while heat treated samples, provided results of 0.128%, 1.42%, and 2.73% NK603. qPCR reactions on sonicated DNA samples, based on the fluorescent dye method, provided results of 0.18%, 0.861% and 1.74% NK603; while heat-treated DNA samples, provided results of 0.18%, 1.02%, and 2.16% NK603. The data suggested that fluorescent dye-based quantifications yielded more accurate determinations of the percent GM-content at higher concentrations most likely because fluorescent dye quantifications provided additional copies of template into the qPCR.
Citation
Food Control
Volume
20
Issue
4

Keywords

A260, degraded DNA quantitative real-time PCR, DNA quantification, Hoescht dye, NK603 maize, Picogreen fluorescence.

Citation

Shokere, L. , Holden, M. and Jenkins, G. (2009), Comparison of Fluorometric and Spectrophotometric DNA Quantification for Real-time Quantitative PCR of Degraded DNA, Food Control (Accessed May 23, 2024)

Issues

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Created March 31, 2009, Updated October 12, 2021