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Single-Molecule Förster Resonance Energy Transfer Methods for Real-Time Investigation of the Holliday Junction Resolution by GEN1
JoVE Journal
Genetics
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JoVE Journal Genetics
Single-Molecule Förster Resonance Energy Transfer Methods for Real-Time Investigation of the Holliday Junction Resolution by GEN1

Single-Molecule Förster Resonance Energy Transfer Methods for Real-Time Investigation of the Holliday Junction Resolution by GEN1

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9,084 Views

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11:27 min

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September 18, 2019

DOI:

10.3791/60045-v

DOI:
10.3791/60045-v

•

11:27 min
September 18, 2019

•

14 Views
, , , , , , , ,
1Laboratory of DNA Replication and Recombination, Biological and Environmental Sciences and Engineering Division (BESE),King Abdullah University of Science and Technology (KAUST)

Summary

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Presented here is a protocol for performing single-molecule Förster resonance energy transfer to study HJ resolution. Two-color alternating excitation is used for determining the dissociation constants. Single-color time lapse smFRET is then applied in real-time cleavage assays to obtain the dwell time distribution prior to HJ resolution.

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Sobhy, M. A., Bralić, A., Raducanu, V., Tehseen, M., Ouyang, Y., Takahashi, M., Rashid, F., Zaher, M. S., Hamdan, S. M. Single-Molecule Förster Resonance Energy Transfer Methods for Real-Time Investigation of the Holliday Junction Resolution by GEN1. J. Vis. Exp. (151), e60045, doi:10.3791/60045 (2019).

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