Strength in Numbers: Development of a Fluorescence Sensor Array for Secondary Structures of DNA.

ABSTRACT

The high-throughput assessment of the secondary structures adopted by DNA oligonucleotides is currently hampered by the lack of suitable biophysical methods. Fluorescent sensors hold great potential in this respect; however, the moderate selectivity that they display for one DNA conformation over the others constitutes a major drawback to the development of accurate assays. Moreover, the use of single sensors impedes a comprehensive classification of the tested sequences. Herein, we propose to overcome these limitations through the development of a fluorescence sensor array constituted by easily accessible, commercial dyes. Multivariate analysis of the emission data matrix produced by the array allows the conformational preferences of DNA sequences of interest to be explored, either by calculating the probability of group membership in the six predefined structural categories (three G-quadruplex groups, double-stranded, and two groups of single-stranded forms) or by revealing their particular structural features. The assay enables rapid screening of synthetic DNA oligonucleotides in a 96-well plate format.