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Single-Molecule Dwell-Time Analysis of Restriction Endonuclease-Mediated DNA Cleavage
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Capitoli
- 00:00Introduction
- 00:31Preparation of Quantum-Dot-Labeled DNA Substrate Materials
- 03:38Microfluidic Device Construction
- 05:58Surface Tethering of Quantum-Dot-Labeled DNA Substrate
- 07:07REase-Mediated DNA Cleavage
- 07:51Results: Single-Molecule TIRF Experiment for Parallelized Observations of DNA Cleavage Events
- 09:32Conclusion
Using quantum-dot-labeled DNA and total internal reflection fluorescence microscopy, we can investigate the reaction mechanism of restriction endonucleases while using unlabeled protein. This single-molecule technique allows for massively multiplexed observation of individual protein-DNA interactions, and data can be pooled to generate well-populated dwell-time distributions.
