Effect of Flow Rate and Ionic Strength on the Stabilities of YOYO-1 and YO-PRO-1 Intercalated in DNA Molecules

Single-molecule DNA studies have improved our understanding of the DNAs' structure and their interactions with other molecules. A variety of DNA labeling dyes are available for single-molecule studies, among which the bis-intercalating dye YOYO-1 and mono-intercalating dye YO-PRO-1 are widely used. They have an extraordinarily strong affinity toward DNA and are bright with a high quantum yield (>0.5) when bound to DNAs. However, it is still not clear how these dyes behave in DNA molecules under higher ionic strength and strong buffer flow. Here, we have studied the effect of ionic strength and flow rate of buffer on their binding in single DNA molecules. The larger the flow rate and the higher the ionic strength, the faster the intercalated dyes are washed away from the DNAs. In the buffer with 1 M ionic strength, YOYO-1 and YO-PRO-1 are mostly washed away from DNA within 2 min of moderate buffer flow.

Automated summary

Single-molecule DNA studies have enhanced our understanding of DNA structure and its interactions with other molecules. YOYO-1 and YO-PRO-1, which are highly-affine and bright DNA labeling dyes, were investigated for their behavior in DNA molecules under high ionic strength and strong buffer flow. It was found that higher flow rate and ionic strength resulted in faster washing away of the intercalated dyes from the DNAs, with YOYO-1 and YO-PRO-1 mostly being washed away within 2 minutes in a buffer with 1 M ionic strength.