UV Decontamination of MDA Reagents for Single Cell Genomics [electronic resource].

Published:: Berkeley, Calif. : Lawrence Berkeley National Laboratory, 2011.
Oak Ridge, Tenn. : Distributed by the Office of Scientific and Technical Information, U.S. Dept. of Energy.
Additional Creators:: Lawrence Berkeley National Laboratory and United States. Department of Energy. Office of Scientific and Technical Information

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Summary:

Single cell genomics, the amplification and sequencing of genomes from single cells, can provide a glimpse into the genetic make-up and thus life style of the vast majority of uncultured microbial cells, making it an immensely powerful and increasingly popular tool. This is accomplished by use of multiple displacement amplification (MDA), which can generate billions of copies of a single bacterial genome producing microgram-range DNA required for shotgun sequencing. Here, we address a key challenge inherent to this approach and propose a solution for the improved recovery of single cell genomes. While DNA-free reagents for the amplification of a single cell genome are a prerequisite for successful single cell sequencing and analysis, DNA contamination has been detected in various reagents, which poses a considerable challenge. Our study demonstrates the effect of UV irradiation in efficient elimination of exogenous contaminant DNA found in MDA reagents, while maintaining Phi29 activity. Consequently, we also find that increased UV exposure to Phi29 does not adversely affect genome coverage of MDA amplified single cells. While additional challenges in single cell genomics remain to be resolved, the proposed methodology is relatively quick and simple and we believe that its application will be of high value for future single cell sequencing projects.

Report Numbers:

E 1.99:lbnl-4514e-poster
lbnl-4514e-poster

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Note:

Published through SciTech Connect.
03/18/2011.
"lbnl-4514e-poster"
DOE JGI User Meeting.
Cheng, Jan-Fang; Woyke, Tanja; Stepanauskas, Ramunas; Malfatti, Stephanie; Wang, Zhong; Rinke, Christian; Lee, Janey; Tighe, Damon; Sczyrba, Alexander; Malmatrom, Rex; Clingenpeel, Scott.
Genomics Division

Funding Information:

DE-AC02-05CH11231

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