A fluorescent intercalator displacement assay for establishing DNA binding selectivity and affinity.
نویسندگان
چکیده
A summary of the qualitative and quantitative elements of a fluorescent intercalator displacement (FID) assay useful for establishing the DNA binding selectivity, affinity, stoichiometry, and binding site size and distinguishing modes of DNA binding is provided.
منابع مشابه
Ligands playing musical chairs with G-quadruplex DNA: a rapid and simple displacement assay for identifying selective G-quadruplex binders.
We report here the details of G4-FID (G-quadruplex fluorescent intercalator displacement), a simple method aiming at evaluating quadruplex-DNA binding affinity and quadruplex- over duplex-DNA selectivity of putative ligands. This assay is based on the loss of fluorescence upon displacement of thiazole orange from quadruplex- and duplex-DNA matrices. The original protocol was tested using variou...
متن کاملFluorescent intercalator displacement replacement (FIDR) assay: determination of relative thermodynamic and kinetic parameters in triplex formation—a case study using triplex-forming LNAs
Triplex forming oligonucleotides (TFOs) are the most commonly used approach for site-specific targeting of double stranded DNA (dsDNA). Important parameters describing triplex formation include equilibrium binding constants (K(eq)) and association/dissociation rate constants (k(on) and k(off)). The 'fluorescent intercalator displacement replacement' (FIDR) assay is introduced herein as an opera...
متن کاملSmall molecule ligands for bulged RNA secondary structures.
A class of wedge-shaped small molecules has been designed, synthesized, and shown to bind bulged RNA secondary structures. These minimally cationic ligands exhibit good affinity and selectivity for certain RNA bulges as demonstrated in a fluorescent intercalator displacement assay.
متن کاملLabel-free detection of kanamycin based on a G-quadruplex DNA aptamer-based fluorescent intercalator displacement assay
This work was the first to report that the kanamycin-binding DNA aptamer (5'-TGG GGG TTG AGG CTA AGC CGA-3') can form stable parallel G-quadruplex DNA (G4-DNA) structures by themselves and that this phenomenon can be verified by nondenaturing polyacrylamide gel electrophoresis and circular dichroism spectroscopy. Based on these findings, we developed a novel label-free strategy for kanamycin de...
متن کاملIdentification of new DNA i-motif binding ligands through a fluorescent intercalator displacement assay† †Electronic supplementary information (ESI) available: Experimental, supporting fluorescence and UV-Vis data, Job plot, binding curves, CD spectra, melting temperatures and SPR data. See DOI: 10.1039/c7ob00710h
i-Motifs are quadruplex DNA structures formed from sequences rich in cytosine and held together by intercalated, hemi-protonated cytosine-cytosine base pairs. These sequences are prevalent in gene promoter regions and may play a role in gene transcription. Targeting these structures with ligands could provide a novel way to target genetic disease but there are very few ligands which have been s...
متن کاملذخیره در منابع من
با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید
برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید
ثبت ناماگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید
ورودعنوان ژورنال:
- Current protocols in nucleic acid chemistry
دوره Chapter 8 شماره
صفحات -
تاریخ انتشار 2004