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Chloroplast sequence variation and the efficacy of peptide nucleic acids for blocking host amplification in plant microbiome studies

Overview of attention for article published in Microbiome, August 2018
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  • In the top 25% of all research outputs scored by Altmetric
  • High Attention Score compared to outputs of the same age (83rd percentile)

Mentioned by

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28 tweeters

Citations

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22 Dimensions

Readers on

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64 Mendeley
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Title
Chloroplast sequence variation and the efficacy of peptide nucleic acids for blocking host amplification in plant microbiome studies
Published in
Microbiome, August 2018
DOI 10.1186/s40168-018-0534-0
Pubmed ID
Authors

Connor R. Fitzpatrick, Patricia Lu-Irving, Julia Copeland, David S. Guttman, Pauline W. Wang, David A. Baltrus, Katrina M. Dlugosch, Marc T. J. Johnson

Abstract

The ability to efficiently characterize microbial communities from host individuals can be limited by co-amplification of host organellar sequences (mitochondrial and/or plastid), which share a common ancestor and thus sequence similarity with extant bacterial lineages. One promising approach is the use of sequence-specific peptide nucleic acid (PNA) clamps, which bind to, and block amplification of, host-derived DNA. Universal PNA clamps have been proposed to block host plant-derived mitochondrial (mPNA) and plastid (pPNA) sequences at the V4 16S rRNA locus, but their efficacy across a wide range of host plant species has not been experimentally tested. Using the universal PNA clamps, we amplified and sequenced root microbial communities from replicate individuals of 32 plant species with a most recent common ancestor inferred at 140 MYA. We found the average rate of host plastid contamination across plant species was 23%, however, particular lineages exhibited much higher rates (62-94%), with the highest levels of contamination occurring in the Asteraceae. We investigated chloroplast sequence variation at the V4 locus across 500 land plant species (Embryophyta) and found six lineages with mismatches between plastid and the universal pPNA sequence, including all species within the Asteraceae. Using a modified pPNA for the Asteraceae sequence, we found (1) host contamination in Asteraceae species was reduced from 65 to 23%; and (2) host contamination in non-Asteraceae species was increased from 12 to 69%. These results demonstrate that even single nucleotide mismatches can lead to drastic reductions in pPNA efficacy in blocking host amplification. Importantly, we found that pPNA type (universal or modified) had no effect on the detection of individual bacterial taxa, or estimates of within and between sample bacterial diversity, suggesting that our modification did not introduce bias against particular bacterial lineages. When high similarity exists between host organellar DNA and PCR target sequences, PNA clamps are an important molecular tool to reduce host contamination during amplification. Here, we provide a validated framework to modify universal PNA clamps to accommodate host variation in organellar sequences.

Twitter Demographics

The data shown below were collected from the profiles of 28 tweeters who shared this research output. Click here to find out more about how the information was compiled.

Mendeley readers

The data shown below were compiled from readership statistics for 64 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Unknown 64 100%

Demographic breakdown

Readers by professional status Count As %
Student > Ph. D. Student 19 30%
Researcher 12 19%
Student > Bachelor 8 13%
Student > Doctoral Student 6 9%
Professor > Associate Professor 3 5%
Other 5 8%
Unknown 11 17%
Readers by discipline Count As %
Agricultural and Biological Sciences 27 42%
Environmental Science 11 17%
Biochemistry, Genetics and Molecular Biology 7 11%
Immunology and Microbiology 2 3%
Engineering 2 3%
Other 4 6%
Unknown 11 17%

Attention Score in Context

This research output has an Altmetric Attention Score of 12. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 25 August 2018.
All research outputs
#1,690,186
of 15,922,891 outputs
Outputs from Microbiome
#630
of 918 outputs
Outputs of similar age
#46,098
of 279,817 outputs
Outputs of similar age from Microbiome
#1
of 1 outputs
Altmetric has tracked 15,922,891 research outputs across all sources so far. Compared to these this one has done well and is in the 89th percentile: it's in the top 25% of all research outputs ever tracked by Altmetric.
So far Altmetric has tracked 918 research outputs from this source. They typically receive a lot more attention than average, with a mean Attention Score of 39.0. This one is in the 31st percentile – i.e., 31% of its peers scored the same or lower than it.
Older research outputs will score higher simply because they've had more time to accumulate mentions. To account for age we can compare this Altmetric Attention Score to the 279,817 tracked outputs that were published within six weeks on either side of this one in any source. This one has done well, scoring higher than 83% of its contemporaries.
We're also able to compare this research output to 1 others from the same source and published within six weeks on either side of this one. This one has scored higher than all of them