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Methods

Evaluation of the Ion Torrent Personal Genome Machine for Gene-Targeted Studies Using Amplicons of the Nitrogenase Gene nifH

Bangzhou Zhang, C. Ryan Penton, Chao Xue, Qiong Wang, Tianling Zheng, James M. Tiedje
C. R. Lovell, Editor
Bangzhou Zhang
aCenter for Microbial Ecology, Michigan State University, East Lansing, Michigan, USA
bState Key Lab of Marine Environmental Science and Key Lab of the MOE for Coastal and Wetland Ecosystems, School of Life Sciences, Xiamen University, Xiamen, China
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C. Ryan Penton
aCenter for Microbial Ecology, Michigan State University, East Lansing, Michigan, USA
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Chao Xue
aCenter for Microbial Ecology, Michigan State University, East Lansing, Michigan, USA
cJiangsu Collaborative Innovation Center for Solid Organic Waste Utilization and National Engineering Research Center for Organic-Based Fertilizers, Department of Plant Nutrition, Nanjing Agricultural University, Nanjing, China
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Qiong Wang
aCenter for Microbial Ecology, Michigan State University, East Lansing, Michigan, USA
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Tianling Zheng
bState Key Lab of Marine Environmental Science and Key Lab of the MOE for Coastal and Wetland Ecosystems, School of Life Sciences, Xiamen University, Xiamen, China
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James M. Tiedje
aCenter for Microbial Ecology, Michigan State University, East Lansing, Michigan, USA
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C. R. Lovell
Roles: Editor
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DOI: 10.1128/AEM.00111-15
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ABSTRACT

The sequencing chips and kits of the Ion Torrent Personal Genome Machine (PGM), which employs semiconductor technology to measure pH changes in polymerization events, have recently been upgraded. The quality of PGM sequences has not been reassessed, and results have not been compared in the context of a gene-targeted microbial ecology study. To address this, we compared sequence profiles across available PGM chips and chemistries and with 454 pyrosequencing data by determining error types and rates and diazotrophic community structures. The PGM was then used to assess differences in nifH-harboring bacterial community structure among four corn-based cropping systems. Using our suggested filters from mock community analyses, the overall error rates were 0.62, 0.36, and 0.39% per base for chips 318 and 314 with the 400-bp kit and chip 318 with the Hi-Q chemistry, respectively. Compared with the 400-bp kit, the Hi-Q kit reduced indel rates by 28 to 59% and produced one to seven times more reads acceptable for downstream analyses. The PGM produced higher frameshift rates than pyrosequencing that were corrected by the RDP FrameBot tool. Significant differences among platforms were identified, although the diversity indices and overall site-based conclusions remained similar. For the cropping system analyses, a total of 6,182 unique NifH operational taxonomic units at 5% amino acid dissimilarity were obtained. The current crop type, as well as the crop rotation history, significantly influenced the composition of the soil diazotrophic community detected.

FOOTNOTES

    • Received 15 January 2015.
    • Accepted 21 April 2015.
    • Accepted manuscript posted online 24 April 2015.
  • Supplemental material for this article may be found at http://dx.doi.org/10.1128/AEM.00111-15.

  • Copyright © 2015, American Society for Microbiology. All Rights Reserved.
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Evaluation of the Ion Torrent Personal Genome Machine for Gene-Targeted Studies Using Amplicons of the Nitrogenase Gene nifH
Bangzhou Zhang, C. Ryan Penton, Chao Xue, Qiong Wang, Tianling Zheng, James M. Tiedje
Applied and Environmental Microbiology Jun 2015, 81 (13) 4536-4545; DOI: 10.1128/AEM.00111-15

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Evaluation of the Ion Torrent Personal Genome Machine for Gene-Targeted Studies Using Amplicons of the Nitrogenase Gene nifH
Bangzhou Zhang, C. Ryan Penton, Chao Xue, Qiong Wang, Tianling Zheng, James M. Tiedje
Applied and Environmental Microbiology Jun 2015, 81 (13) 4536-4545; DOI: 10.1128/AEM.00111-15
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