Exploring interaction of dye with DNA bases
Dy630, Dy632, and Dy635 are red fluorescent dyes which are used routinely in labeling biomolecules. This study focuses on seeing how cis-trans isomerization is affected by changing the environment of the dye, i.e., by placing it on a hairpin forming ssDNA, and dsDNA. Temperature dependent fluorescence change was used to calculate activation energy of conformation change (trans to cis) for different systems. deoxynucleoside monophosphate (dNMP) based fluorescence change is being studied to see how different natural bases interact with the dye.
Increase in fluorescence intensity observed when a protein binds to a nucleic acid in the proximity of a fluorescent probe is termed as protein induced fluorescence enhancement (PIFE). The next step for us is to see if this dye exhibits PIFE using single molecular experiment. For this, we have built a home-made total internal reflection fluorescence (TIRF) microscope. Total internal reflection is used here to excite the fluorophores on the quartz surface which are immobilized by avidin-biotin strategy. Work is going to optimize the parameters for the setup and use it to observe PIFE with dy dyes.