Investigation of various fluorescent protein–DNA binding peptides for effectively visualizing large DNA molecules

Abstract

Single-molecule DNA visualization with fluorescent protein DNA binding peptides.

TAMRA-polypyrrole for A/T sequence visualization on DNA molecules

“…This predicted value agrees relatively well with 14.03 μm (86%) observed for TAMRA-polypyrrole stained λ DNA in this report ( Supplementary Figure S3 ). In addition, this stretching result is consistent with stretching values (13.4–14.6 μm) for various fluorescent DNA binding proteins to stain DNA molecules in the same flow cell that we reported earlier ( 32 ). Therefore, 86% DNA stretch suggests that TAMRA-polypyrrole does not cause considerable structural deformation ( Supplementary Figure S3 ).…”

“…Therefore, the DNA stretching rates indicated the degree of dye-induced structural deformation such as double helix unwinding. In our previous study, we reported that YOYO-1-stained DNA exhibited severe structural deformations resulting in 21.8 μm λ DNA in a flow cell, which corresponds to 134% stretching compared with native λ DNA (21.8/16.3 μm), but our previous computer simulation using finite element analysis predicted that native DNA should stretch 84% in the same flow cell ( 32 ). This predicted value agrees relatively well with 14.03 μm (86%) observed for TAMRA-polypyrrole stained λ DNA in this report ( Supplementary Figure S3 ).…”

Nanochannel-Confined TAMRA-Polypyrrole Stained DNA Stretching by Varying the Ionic Strength from Micromolar to Millimolar Concentrations

“…First of all, TAMRA-polypyrrole stained DNA ( 2 in Figure 1) stretched up to 14.03 ± 0.54 µm ( X / L = 86% ± 3%) at 100 µL/min within 100 µm high flow cell [29]. Previously, we calculated the force of 0.79 pN applied to the λ DNA in the same flow cell (100 µL/min) [27]. Based on this value, we checked Bustamante’s paper, in which they measured DNA stretches versus the force by optical tweezers.…”

“…However, it is notable that this stretch would be 88% ± 3% compared with Murade’s contour length (24.67 µm) for YOYO-1 stained DNA [28]. For comparison, we added DNA stretches stained by FP-DBPs (DNA binding peptides) from 6 to 10 in Figure 1 [21,27]. Figure 1 suggests that TAMRA-polypyrrole stained DNA preserves the contour length of natural B-form DNA, while YOYO-1 stained DNA changes it considerably.…”

“…Recently, we have developed several novel DNA staining reagents, such as fluorescent protein-DNA binding peptides (FP-DBPs) [21,27,28] and TAMRA-polypyrrole [29]. They consist of a fluorophore and a DNA binding moiety, which makes the fluorophore well separated from the DNA backbone.…”