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Microbial Environmental Genomics (MEG)

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Cover of 'Microbial Environmental Genomics (MEG)'

Table of Contents

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    Book Overview
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    Chapter 1 "Deciphering Archaeal Communities" Omics Tools in the Study of Archaeal Communities.
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    Chapter 2 Investigating the Endobacteria Which Thrive in Arbuscular Mycorrhizal Fungi
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    Chapter 3 GenoSol Platform: A Logistic and Technical Platform for Conserving and Exploring Soil Microbial Diversity.
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    Chapter 4 Sample Preparation for Fungal Community Analysis by High-Throughput Sequencing of Barcode Amplicons
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    Chapter 5 Fungal Communities in Soils: Soil Organic Matter Degradation.
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    Chapter 6 DNA-Based Characterization and Identification of Arbuscular Mycorrhizal Fungi Species.
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    Chapter 7 Molecular Identification of Soil Eukaryotes and Focused Approaches Targeting Protist and Faunal Groups Using High-Throughput Metabarcoding.
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    Chapter 8 Identification and In Situ Distribution of a Fungal Gene Marker: The Mating Type Genes of the Black Truffle
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    Chapter 9 Stable-Isotope Probing RNA to Study Plant/Fungus Interactions
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    Chapter 10 Targeted Gene Capture by Hybridization to Illuminate Ecosystem Functioning.
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    Chapter 11 Hybridization of Environmental Microbial Community Nucleic Acids by GeoChip.
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    Chapter 12 Reconstruction of Transformation Processes Catalyzed by the Soil Microbiome Using Metagenomic Approaches.
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    Chapter 13 MG-RAST, a Metagenomics Service for Analysis of Microbial Community Structure and Function.
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    Chapter 14 Analysis of Active Methylotrophic Communities: When DNA-SIP Meets High-Throughput Technologies.
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    Chapter 15 Functional Metagenomics: Construction and High-Throughput Screening of Fosmid Libraries for Discovery of Novel Carbohydrate-Active Enzymes.
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    Chapter 16 Metatranscriptomics of Soil Eukaryotic Communities
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    Chapter 17 Analysis of Ancient DNA in Microbial Ecology.
Attention for Chapter 4: Sample Preparation for Fungal Community Analysis by High-Throughput Sequencing of Barcode Amplicons
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Chapter title
Sample Preparation for Fungal Community Analysis by High-Throughput Sequencing of Barcode Amplicons
Chapter number 4
Book title
Microbial Environmental Genomics (MEG)
Published in
Methods in molecular biology, January 2016
DOI 10.1007/978-1-4939-3369-3_4
Pubmed ID
Book ISBNs
978-1-4939-3367-9, 978-1-4939-3369-3
Authors

Karina Engelbrecht Clemmensen, Katarina Ihrmark, Mikael Brandström Durling, Björn D. Lindahl

Editors

Francis Martin, Stephane Uroz

Abstract

Fungal species participate in vast numbers of processes in the landscape around us. However, their often cryptic growth, inside various substrates and in highly diverse species assemblages, has been a major obstacle to thorough analysis of fungal communities, hampering exhaustive description of the fungal kingdom. Recent technological developments allowing rapid, high-throughput sequencing of mixed communities from many samples at once are currently having a tremendous impact in fungal community ecology. Universal DNA extraction followed by amplification and sequencing of fungal species-level barcodes such as the nuclear internal transcribed spacer (ITS) region now enable identification and relative quantification of fungal community members across well-replicated experimental settings.Here, we present the sample preparation procedure presently used in our laboratory for fungal community analysis by high-throughput sequencing of amplified ITS2 markers. We focus on the procedure optimized for studies of total fungal communities in humus-rich soils, wood, and litter. However, this procedure can be applied to other sample types and markers. We focus on the laboratory-based part of sample preparation, that is, the procedure from the point where samples enter the laboratory until amplicons are submitted for sequencing. Our procedure comprises four main parts: (1) universal DNA extraction, (2) optimization of PCR conditions, (3) production of tagged ITS amplicons, and (4) preparation of the multiplexed amplicon mix to be sequenced. The presented procedure is independent of the specific high-throughput sequencing technology used, which makes it highly versatile.

Mendeley readers

The data shown below were compiled from readership statistics for 32 Mendeley readers of this research output. Click here to see the associated Mendeley record.

Geographical breakdown

Country Count As %
Unknown 32 100%

Demographic breakdown

Readers by professional status Count As %
Researcher 7 22%
Student > Master 7 22%
Student > Ph. D. Student 6 19%
Student > Doctoral Student 3 9%
Professor > Associate Professor 2 6%
Other 3 9%
Unknown 4 13%
Readers by discipline Count As %
Agricultural and Biological Sciences 15 47%
Environmental Science 5 16%
Biochemistry, Genetics and Molecular Biology 4 13%
Veterinary Science and Veterinary Medicine 1 3%
Earth and Planetary Sciences 1 3%
Other 2 6%
Unknown 4 13%