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Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi

Arbuscular mycorrhiza fungi (AMF) are beneficial soil fungi that can promote the growth of their host plants. Accurate quantification of AMF in plant roots is important because the level of colonization is often indicative of the activity of these fungi. Root colonization is traditionally measured w...

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Autores principales: Bodenhausen, Natacha, Deslandes-Hérold, Gabriel, Waelchli, Jan, Held, Alain, van der Heijden, Marcel G. A., Schlaeppi, Klaus
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Springer Berlin Heidelberg 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7910240/
https://www.ncbi.nlm.nih.gov/pubmed/33475800
http://dx.doi.org/10.1007/s00572-020-01014-1
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author Bodenhausen, Natacha
Deslandes-Hérold, Gabriel
Waelchli, Jan
Held, Alain
van der Heijden, Marcel G. A.
Schlaeppi, Klaus
author_facet Bodenhausen, Natacha
Deslandes-Hérold, Gabriel
Waelchli, Jan
Held, Alain
van der Heijden, Marcel G. A.
Schlaeppi, Klaus
author_sort Bodenhausen, Natacha
collection PubMed
description Arbuscular mycorrhiza fungi (AMF) are beneficial soil fungi that can promote the growth of their host plants. Accurate quantification of AMF in plant roots is important because the level of colonization is often indicative of the activity of these fungi. Root colonization is traditionally measured with microscopy methods which visualize fungal structures inside roots. Microscopy methods are labor-intensive, and results depend on the observer. In this study, we present a relative qPCR method to quantify AMF in which we normalized the AMF qPCR signal relative to a plant gene. First, we validated the primer pair AMG1F and AM1 in silico, and we show that these primers cover most AMF species present in plant roots without amplifying host DNA. Next, we compared the relative qPCR method with traditional microscopy based on a greenhouse experiment with Petunia plants that ranged from very high to very low levels of AMF root colonization. Finally, by sequencing the qPCR amplicons with MiSeq, we experimentally confirmed that the primer pair excludes plant DNA while amplifying mostly AMF. Most importantly, our relative qPCR approach was capable of discriminating quantitative differences in AMF root colonization and it strongly correlated (Spearman Rho = 0.875) with quantifications by traditional microscopy. Finally, we provide a balanced discussion about the strengths and weaknesses of microscopy and qPCR methods. In conclusion, the tested approach of relative qPCR presents a reliable alternative method to quantify AMF root colonization that is less operator-dependent than traditional microscopy and offers scalability to high-throughput analyses.
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spelling pubmed-79102402021-03-15 Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi Bodenhausen, Natacha Deslandes-Hérold, Gabriel Waelchli, Jan Held, Alain van der Heijden, Marcel G. A. Schlaeppi, Klaus Mycorrhiza Original Article Arbuscular mycorrhiza fungi (AMF) are beneficial soil fungi that can promote the growth of their host plants. Accurate quantification of AMF in plant roots is important because the level of colonization is often indicative of the activity of these fungi. Root colonization is traditionally measured with microscopy methods which visualize fungal structures inside roots. Microscopy methods are labor-intensive, and results depend on the observer. In this study, we present a relative qPCR method to quantify AMF in which we normalized the AMF qPCR signal relative to a plant gene. First, we validated the primer pair AMG1F and AM1 in silico, and we show that these primers cover most AMF species present in plant roots without amplifying host DNA. Next, we compared the relative qPCR method with traditional microscopy based on a greenhouse experiment with Petunia plants that ranged from very high to very low levels of AMF root colonization. Finally, by sequencing the qPCR amplicons with MiSeq, we experimentally confirmed that the primer pair excludes plant DNA while amplifying mostly AMF. Most importantly, our relative qPCR approach was capable of discriminating quantitative differences in AMF root colonization and it strongly correlated (Spearman Rho = 0.875) with quantifications by traditional microscopy. Finally, we provide a balanced discussion about the strengths and weaknesses of microscopy and qPCR methods. In conclusion, the tested approach of relative qPCR presents a reliable alternative method to quantify AMF root colonization that is less operator-dependent than traditional microscopy and offers scalability to high-throughput analyses. Springer Berlin Heidelberg 2021-01-21 2021 /pmc/articles/PMC7910240/ /pubmed/33475800 http://dx.doi.org/10.1007/s00572-020-01014-1 Text en © The Author(s) 2021 Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.
spellingShingle Original Article
Bodenhausen, Natacha
Deslandes-Hérold, Gabriel
Waelchli, Jan
Held, Alain
van der Heijden, Marcel G. A.
Schlaeppi, Klaus
Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi
title Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi
title_full Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi
title_fullStr Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi
title_full_unstemmed Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi
title_short Relative qPCR to quantify colonization of plant roots by arbuscular mycorrhizal fungi
title_sort relative qpcr to quantify colonization of plant roots by arbuscular mycorrhizal fungi
topic Original Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7910240/
https://www.ncbi.nlm.nih.gov/pubmed/33475800
http://dx.doi.org/10.1007/s00572-020-01014-1
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