Abstract Fluorescent nucleoside analogues with strong and informative responses to their local environment are in urgent need for DNA research. In this work, the design, synthesis and investigation of a new solvatochromic ratiometric fluorophore compiled from 3- hydroxychromones (3HCs) and uracil fragments are reported. 3HC dyes are a class of multi- parametric, environment-sensitive fluorophores providing a ratiometric response due to ...