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High Throughput, Low Volume DNA Quantitation Using the QuantiFluor dsDNA System

Promega Corporation
Publication Date: 9/17

Abstract

DNA quantitation and normalization are critical steps for a wide range of molecular biology applications such as cloning, quantitative PCR (qPCR) and next-generation sequencing (NGS). Fluorescent dye-based DNA quantitation uses specially designed DNA binding compounds that intercalate only with double-stranded DNA (dsDNA) molecules. When excited by a specific wavelength of light, only dye in the DNA-bound state will fluoresce. These aspects of the technique contribute to low background signal, high accuracy and specificity—making it a suitable measure of DNA concentration prior to advanced downstream applications. Another desirable attribute of fluorescence quantitation over other methods is ease of use and scalability for laboratories conducting all levels of throughput. In this application note, we demonstrate the flexibility of the QuantiFluor® dsDNA System for use in 40μl reaction volumes compatible with 384-well plates. We also provide guidance for optimization of the assay according to variables such as dynamic range, sensitivity, accuracy, cost per reaction and other features of high-throughput workflows.

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