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PMID: 22028825 Published · ppublish English Journal Article Research Support, U.S. Gov't, Non-P.H.S.

Decontamination of MDA reagents for single cell whole genome amplification.

PloS one ·Vol. 6 ·No. 10 ·2011-00-00 ·Pages e26161

Woyke T, Sczyrba A, Lee J, Rinke C, Tighe D, Clingenpeel S, Malmstrom R, Stepanauskas R, Cheng JF

Abstract

Single cell genomics is a powerful and increasingly popular tool for studying the genetic make-up of uncultured microbes. A key challenge for successful single cell sequencing and analysis is the removal of exogenous DNA from whole genome amplification reagents. We found that UV irradiation of the multiple displacement amplification (MDA) reagents, including the Phi29 polymerase and random hexamer primers, effectively eliminates the amplification of contaminating DNA. The methodology is quick, simple, and highly effective, thus significantly improving whole genome amplification from single cells.

MeSH Terms
Artifacts Bacillus Phages/enzymology DNA Contamination DNA Primers/genetics,metabolism DNA-Directed DNA Polymerase/metabolism Dose-Response Relationship, Radiation Escherichia coli K12/cytology,genetics Genomics/methods Indicators and Reagents/metabolism Nucleic Acid Amplification Techniques/methods Sequence Analysis, DNA Single-Cell Analysis/methods Time Factors Ultraviolet Rays
Chemicals
DNA Primers Indicators and Reagents DNA-Directed DNA Polymerase
Authors & Affiliations
9 authors, click to expand affiliations / ORCID
Woyke Tanja
Department of Energy Joint Genome Institute, Walnut Creek, California, United States of America.
Sczyrba Alexander
Lee Janey
Rinke Christian
Tighe Damon
Clingenpeel Scott
Malmstrom Rex
Stepanauskas Ramunas
Cheng Jan-Fang
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Article Info
Journal
PloS one
Abbr.
PLoS One
ISSN
1932-6203
Published
2011-00-00
Epub
2011-00-20
Pages
e26161
Language
English
Region
United States
NLM ID
101285081
PMCID
PMC3197606
Subset
IM
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