ArticleBMC microbiology2015
Phasing amplicon sequencing on Illumina Miseq for robust environmental microbial community analysis.
Article in BMC microbiology, 2015. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 133 papers.
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133 citing papers in PubMed.
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- Increased precipitation decelerates temporal succession of grassland soil microbial communities.The ISME journal · 2026Article
- The Appalbees menu: a multiyear, multilocus metagenetic assessment of pollen foraging by AppalachianPeerJ · 2026Article
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- De Novo Design of Miniprotein Inhibitors of Bacterial Adhesins.Research square · 2025Article
- Nontuberculous mycobacteria remodel lung microbiota in cystic fibrosis-associated respiratory infections.Microbiology spectrum · 2025Article
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- Tracing non-fungal eukaryotic diversity via shotgun metagenomes in the complex mudflat intertidal zones.mSystems · 2025Article
- Feeding Behavior and Ecological Significance ofBiology · 2025Article
- Data of the Insect Biome Atlas: a metabarcoding survey of the terrestrial arthropods of Sweden and Madagascar.Scientific data · 2025Article
- Altered precipitation and nighttime warming reshape the vertical distribution of soil microbial communities.mSystems · 2025Article
- Modest functional diversity decline and pronounced composition shifts of microbial communities in a mixed waste-contaminated aquifer.Microbiome · 2025Article
- Microfluidic droplets with amended culture media cultivate a greater diversity of soil microorganisms.Applied and environmental microbiology · 2025Article
73 more citing papers are in PubMed but not listed here.
Corrections and comments
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Authors and funding
10 authors.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundAlthough high-throughput sequencing, such as Illumina-based technologies (e.g. MiSeq), has revolutionized microbial ecology, adaptation of amplicon sequencing for environmental microbial community analysis is challenging due to the problem of low base diversity.
resultsA new phasing amplicon sequencing approach (PAS) was developed by shifting sequencing phases among different community samples from both directions via adding various numbers of bases (0-7) as spacers to both forward and reverse primers. Our results first indicated that the PAS method substantially ameliorated the problem of unbalanced base composition. Second, the PAS method substantially improved the sequence read base quality (an average of 10 % higher of bases above Q30). Third, the PAS method effectively increased raw sequence throughput (~15 % more raw reads). In addition, the PAS method significantly increased effective reads (9-47 %) and the effective read sequence length (16-96 more bases) after quality trim at Q30 with window 5. In addition, the PAS method reduced half of the sequencing errors (0.54-1.1 % less). Finally, two-step PCR amplification of the PAS method effectively ameliorated the amplification biases introduced by the long barcoded PCR primers.
conclusionThe developed strategy is robust for 16S rRNA gene amplicon sequencing. In addition, a similar strategy could also be used for sequencing other genes important to ecosystem functional processes.
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