The DNA–protein interaction kit can kick-start your single-molecule experiments, allowing you to study the kinetics of enzymatic activities associated with DNA replication. The biotinylated pKYB1 DNA (8,398 bp) contains a 25-nucleotide long 5′-overhang that serves as a loading site for DNA polymerases. This kit is ideal for extracting real-time data of DNA replication processes, including polymerization dynamics, proofreading activities, and pause states.
The DNA–protein interactions kit provides you with a ready-to-go system for single-molecule applications to:
- Obtain millisecond and base-pair resolution data with kinetics and mechanistic information from DNA-replication processes at the single-molecule level.
- Mimic nucleotide miss-incorporation through the application of DNA tension forces, which enables you to measure polymerization rates, exonucleolytic activity, and pause states of DNA polymerase enzymes.
- Test polymerase activity with the u-Flux™ Laminar Flow Microfluidics, which facilitates switching between different experimental conditions in different channels.
T7 DNA polymerase
20 µl, 200 Units – 10000 U/ml (typical dilution factor 1:500) T7 DNA polymerase with 8x Histidine tag. In 50% glycerol.
Biotinylated double-stranded pKYB1 DNA
20 µl – 20 ng/µl (typical dilution factor 1:1000) double-stranded expression vector pKYB1 (8393 bp) with a 25-nt 5′-end overhang. Biotin-labeled on the 5′ and 3′ ends of one of the strands, diluted in TE buffer (10 mM Tris HCl pH 8.0 and 1 mM EDTA).
300 µl – 10 mM of each (typical dilution factor 1:20) deoxynucleotide (dNTP) mix with equimolar concentration dATP, dCTP, dGTP and dTTP. Used by T7 DNA polymerase for incorporating nucleotides.
500 µl – (typical dilution factor 1:20) magnesium based buffer to catalyze the activity of T7 DNA polymerase.
Polystyrene beads (1.76 µm)
25 µl – 1% (w/v) (typical dilution factor 1:1000) streptavidin coated polystyrene microspheres with a diameter of 1.76 µm in water. Binds to biotinylated DNA (e.g., the provided pkYB1) and can be optically manipulated using your optical tweezers system.
Want to learn more?
See our DNA/RNA–Protein Interactions application notes.
Want to use this product in practice?
Find out more about the C-Trap here.