Multiplexed single-molecule assay for enzymatic activity on flow-stretched DNA

Sangjin Kim; Paul C. Blainey; Charles M. Schroeder; X. Sunney Xie

doi:10.1038/nmeth1037

Multiplexed single-molecule assay for enzymatic activity on flow-stretched DNA

Sangjin Kim, Paul C. Blainey, Charles M. Schroeder, X. Sunney Xie

Research output: Contribution to journal › Article › peer-review

Abstract

We report a single-molecule assay for nucleic-acid enzymes on flow-stretched DNA templates. To facilitate the detection of slow or intermittent enzymatic activities, we developed the assay with 15-nm spatial resolution at a frame rate of 1 Hz and ∼10 nm mechanical stability over the timescale of hours. With multiplexed data collection, we applied the assay to φ29 DNA polymerase, HIV-1 reverse transcriptase, λ exonuclease and Escherichia coli RNA polymerase.

Original language	English (US)
Pages (from-to)	397-399
Number of pages	3
Journal	Nature Methods
Volume	4
Issue number	5
DOIs	https://doi.org/10.1038/nmeth1037
State	Published - May 2007
Externally published	Yes

ASJC Scopus subject areas

Biotechnology
Biochemistry
Molecular Biology
Cell Biology

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10.1038/nmeth1037

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title = "Multiplexed single-molecule assay for enzymatic activity on flow-stretched DNA",

abstract = "We report a single-molecule assay for nucleic-acid enzymes on flow-stretched DNA templates. To facilitate the detection of slow or intermittent enzymatic activities, we developed the assay with 15-nm spatial resolution at a frame rate of 1 Hz and ∼10 nm mechanical stability over the timescale of hours. With multiplexed data collection, we applied the assay to φ29 DNA polymerase, HIV-1 reverse transcriptase, λ exonuclease and Escherichia coli RNA polymerase.",

author = "Sangjin Kim and Blainey, {Paul C.} and Schroeder, {Charles M.} and Xie, {X. Sunney}",

note = "Funding Information: We thank A. van Oijen for his initial contribution to the flow-stretching assay and H. Babcock for suggesting a pressure-driven pump. This work is supported by Jane Coffin Childs Memorial Fund for Medical Research Fellowship and a US National Institutes of Health Pathway to Independence Award for C.M.S. and a National Institutes of Health Director{\textquoteright}s Pioneer Award to X.S.X.",

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Multiplexed single-molecule assay for enzymatic activity on flow-stretched DNA

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