Quantifying double-stranded DNA with fluorescent dyes: Hoechst 33258 measured on a FLUOstar Omega
Desoxyribonucleic acid (DNA) is the storage medium of genetic information making it a major subject of basic research and diagnostics. Nowadays, so-called next generation sequencing (NGS) methods uncover base sequences and mutations in hours and at reasonable costs. Prerequisite for reliable sequencing data is accurate preparation and quantitation of the DNA library. Fluorescent dsDNA dyes are frequently used to quantify dsDNA as they offer high sensitivity and specificity.
Here, we present DNA quantification by the cost-effective Hoechst 33258 DNA binding fluorophore. Its detection on a FLUOstar® Omega resulted in high linearity and precision for measuring 11-300 ng/well dsDNA. The method is compatible with high throughput and can be fully automated when integrating the microplate reader into a liquid handling system. This facilitates the integration of DNA quantification into an automated NGS workflow.