PMID- 34546069
OWN - NLM
STAT- PubMed-not-MEDLINE
LR  - 20211028
IS  - 2379-5077 (Print)
IS  - 2379-5077 (Electronic)
IS  - 2379-5077 (Linking)
VI  - 6
IP  - 5
DP  - 2021 Oct 26
TI  - MetaFunPrimer: an Environment-Specific, High-Throughput Primer Design Tool for 
      Improved Quantification of Target Genes.
PG  - e0020121
LID - 10.1128/mSystems.00201-21 [doi]
LID - e00201-21
AB  - Genes belonging to the same functional group may include numerous and variable 
      gene sequences, making characterizing and quantifying difficult. Therefore, 
      high-throughput design tools are needed to simultaneously create primers for 
      improved quantification of target genes. We developed MetaFunPrimer, a 
      bioinformatic pipeline, to design primers for numerous genes of interest. This 
      tool also enables gene target prioritization based on ranking the presence of 
      genes in user-defined references, such as environment-specific metagenomes. Given 
      inputs of protein and nucleotide sequences for gene targets of interest and an 
      accompanying set of reference metagenomes or genomes, MetaFunPrimer generates 
      primers for ranked genes of interest. To demonstrate the usage and benefits of 
      MetaFunPrimer, a total of 78 primer pairs were designed to target observed 
      ammonia monooxygenase subunit A (amoA) genes of ammonia-oxidizing bacteria (AOB) 
      in 1,550 publicly available soil metagenomes. We demonstrate computationally that 
      these amoA-AOB primers can cover 94% of the amoA-AOB genes observed in the 1,550 
      soil metagenomes compared with a 49% estimated coverage by previously published 
      primers. Finally, we verified the utility of these primer sets in incubation 
      experiments that used long-term nitrogen fertilized or unfertilized soils. 
      High-throughput quantitative PCR (qPCR) results and statistical analyses showed 
      significant differences in relative quantification patterns between the two 
      soils, and subsequent absolute quantifications also confirmed that target genes 
      enumerated by six selected primer pairs were significantly more abundant in the 
      nitrogen-fertilized soils. This new tool gives microbial ecologists a new 
      approach to assess functional gene abundance and related microbial community 
      dynamics quickly and affordably. IMPORTANCE Amplification-based gene 
      characterization allows for sensitive and specific quantification of functional 
      genes. There is often a large diversity of genes represented for functional gene 
      groups, and multiple primers may be necessary to target associated genes. Current 
      primer design tools are limited to designing primers for only a few genes of 
      interest. MetaFunPrimer allows for high-throughput primer design for various 
      genes of interest and also allows for ranking gene targets by their presence and 
      abundance in environmental data sets. Primers designed by this tool improve the 
      characterization and quantification of functional genes in broad gene 
      amplification platforms and can be powerful with high-throughput qPCR approaches.
FAU - Liu, Jia
AU  - Liu J
AD  - Department of Agricultural and Biosystems Engineering, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Bioinformatics and Computational Biology Department, Iowa State University, Ames, 
      Iowa, USA.
AD  - Center for Advanced Bioenergy and Bioproducts Innovation, Urbana, Illinois, USA.
FAU - Villanueva, Paul
AU  - Villanueva P
AD  - Department of Agricultural and Biosystems Engineering, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Bioinformatics and Computational Biology Department, Iowa State University, Ames, 
      Iowa, USA.
FAU - Choi, Jinlyung
AU  - Choi J
AD  - Department of Agricultural and Biosystems Engineering, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Center for Advanced Bioenergy and Bioproducts Innovation, Urbana, Illinois, USA.
FAU - Gunturu, Santosh
AU  - Gunturu S
AD  - Center for Microbial Ecology, Michigan State Universitygrid.17088.36, East 
      Lansing, Michigan, USA.
AD  - DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, 
      Madison, Wisconsin, USA.
FAU - Ouyang, Yang
AU  - Ouyang Y
AD  - Department of Plant, Soil, and Microbial Sciences, Michigan State 
      Universitygrid.17088.36, East Lansing, Michigan, USA.
AD  - DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, 
      Madison, Wisconsin, USA.
FAU - Tiemann, Lisa K
AU  - Tiemann LK
AD  - Department of Plant, Soil, and Microbial Sciences, Michigan State 
      Universitygrid.17088.36, East Lansing, Michigan, USA.
AD  - DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, 
      Madison, Wisconsin, USA.
FAU - Cole, James R
AU  - Cole JR
AD  - Department of Plant, Soil, and Microbial Sciences, Michigan State 
      Universitygrid.17088.36, East Lansing, Michigan, USA.
AD  - Center for Microbial Ecology, Michigan State Universitygrid.17088.36, East 
      Lansing, Michigan, USA.
AD  - DOE Great Lakes Bioenergy Research Center, University of Wisconsin-Madison, 
      Madison, Wisconsin, USA.
FAU - Glanville, Kathryn R
AU  - Glanville KR
AD  - Department of Agricultural and Biosystems Engineering, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Center for Advanced Bioenergy and Bioproducts Innovation, Urbana, Illinois, USA.
FAU - Hall, Steven J
AU  - Hall SJ
AD  - Department of Ecology, Evolution, and Organismal Biology, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
FAU - McDaniel, Marshall D
AU  - McDaniel MD
AD  - Department of Agronomy, Iowa State Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Center for Advanced Bioenergy and Bioproducts Innovation, Urbana, Illinois, USA.
FAU - Lee, Jaejin
AU  - Lee J
AD  - Department of Agricultural and Biosystems Engineering, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Center for Advanced Bioenergy and Bioproducts Innovation, Urbana, Illinois, USA.
FAU - Howe, Adina
AU  - Howe A
AUID- ORCID: 0000-0002-7705-343X
AD  - Department of Agricultural and Biosystems Engineering, Iowa State 
      Universitygrid.34421.30, Ames, Iowa, USA.
AD  - Bioinformatics and Computational Biology Department, Iowa State University, Ames, 
      Iowa, USA.
AD  - Center for Advanced Bioenergy and Bioproducts Innovation, Urbana, Illinois, USA.
LA  - eng
GR  - DE-SC0018420/U.S. Department of Energy (DOE)/
GR  - DE-SC0014108/U.S. Department of Energy (DOE)/
PT  - Journal Article
DEP - 20210921
PL  - United States
TA  - mSystems
JT  - mSystems
JID - 101680636
PMC - PMC8547451
OTO - NOTNLM
OT  - environment-specific
OT  - functional gene quantification
OT  - nitrification
OT  - primer design
OT  - quantitative PCR
EDAT- 2021/09/22 06:00
MHDA- 2021/09/22 06:01
PMCR- 2021/09/21
CRDT- 2021/09/21 12:18
PHST- 2021/09/22 06:00 [pubmed]
PHST- 2021/09/22 06:01 [medline]
PHST- 2021/09/21 12:18 [entrez]
PHST- 2021/09/21 00:00 [pmc-release]
AID - mSystems00201-21 [pii]
AID - msystems.00201-21 [pii]
AID - 10.1128/mSystems.00201-21 [doi]
PST - ppublish
SO  - mSystems. 2021 Oct 26;6(5):e0020121. doi: 10.1128/mSystems.00201-21. Epub 2021 
      Sep 21.