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Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae
Clubroot, caused by the soilborne pathogen Plasmodiophora brassicae, is an important disease of canola (Brassica napus) and other crucifers. The recent application of RNA sequencing (RNA-seq) technologies to study P. brassicae–host interactions has generated large amounts of gene expression data, im...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9779335/ https://www.ncbi.nlm.nih.gov/pubmed/36555223 http://dx.doi.org/10.3390/ijms232415581 |
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author | Zhou, Qinqin Galindo-González, Leonardo Hwang, Sheau-Fang Strelkov, Stephen E. |
author_facet | Zhou, Qinqin Galindo-González, Leonardo Hwang, Sheau-Fang Strelkov, Stephen E. |
author_sort | Zhou, Qinqin |
collection | PubMed |
description | Clubroot, caused by the soilborne pathogen Plasmodiophora brassicae, is an important disease of canola (Brassica napus) and other crucifers. The recent application of RNA sequencing (RNA-seq) technologies to study P. brassicae–host interactions has generated large amounts of gene expression data, improving knowledge of the molecular mechanisms of pathogenesis and host resistance. Quantitative PCR (qPCR) analysis has been widely applied to examine the expression of a limited number of genes and to validate the results of RNA-seq studies, but may not be ideal for analyzing larger suites of target genes or increased sample numbers. Moreover, the need for intermediate steps such as cDNA synthesis may introduce variability that could affect the accuracy of the data generated by qPCR. Here, we report the validation of gene expression data from a previous RNA-seq study of clubroot using the NanoString nCounter System, which achieves efficient gene expression quantification in a fast and simple manner. We first confirm the robustness of the NanoString system by comparing the results with those generated by qPCR and RNA-seq and then discuss the importance of some candidate genes for resistance or susceptibility to P. brassicae in the host. The results show that the expression of genes measured using NanoString have a high correlation with the values obtained using the other two technologies, with R > 0.90 and p < 0.01, and the same expression patterns for most genes. The three methods (qPCR, RNA-seq, and NanoString) were also compared in terms of laboratory procedures, time, and cost. We propose that the NanoString nCounter System is a robust, sensitive, highly reproducible, and simple technology for gene expression analysis. NanoString could become a common alternative to qPCR to validate RNA-seq data or to create panels of genes for use as markers of resistance/susceptibility when plants are challenged with different P. brassicae pathotypes. |
format | Online Article Text |
id | pubmed-9779335 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-97793352022-12-23 Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae Zhou, Qinqin Galindo-González, Leonardo Hwang, Sheau-Fang Strelkov, Stephen E. Int J Mol Sci Article Clubroot, caused by the soilborne pathogen Plasmodiophora brassicae, is an important disease of canola (Brassica napus) and other crucifers. The recent application of RNA sequencing (RNA-seq) technologies to study P. brassicae–host interactions has generated large amounts of gene expression data, improving knowledge of the molecular mechanisms of pathogenesis and host resistance. Quantitative PCR (qPCR) analysis has been widely applied to examine the expression of a limited number of genes and to validate the results of RNA-seq studies, but may not be ideal for analyzing larger suites of target genes or increased sample numbers. Moreover, the need for intermediate steps such as cDNA synthesis may introduce variability that could affect the accuracy of the data generated by qPCR. Here, we report the validation of gene expression data from a previous RNA-seq study of clubroot using the NanoString nCounter System, which achieves efficient gene expression quantification in a fast and simple manner. We first confirm the robustness of the NanoString system by comparing the results with those generated by qPCR and RNA-seq and then discuss the importance of some candidate genes for resistance or susceptibility to P. brassicae in the host. The results show that the expression of genes measured using NanoString have a high correlation with the values obtained using the other two technologies, with R > 0.90 and p < 0.01, and the same expression patterns for most genes. The three methods (qPCR, RNA-seq, and NanoString) were also compared in terms of laboratory procedures, time, and cost. We propose that the NanoString nCounter System is a robust, sensitive, highly reproducible, and simple technology for gene expression analysis. NanoString could become a common alternative to qPCR to validate RNA-seq data or to create panels of genes for use as markers of resistance/susceptibility when plants are challenged with different P. brassicae pathotypes. MDPI 2022-12-08 /pmc/articles/PMC9779335/ /pubmed/36555223 http://dx.doi.org/10.3390/ijms232415581 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Zhou, Qinqin Galindo-González, Leonardo Hwang, Sheau-Fang Strelkov, Stephen E. Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae |
title | Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae |
title_full | Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae |
title_fullStr | Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae |
title_full_unstemmed | Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae |
title_short | Application of the NanoString nCounter System as an Alternative Method to Investigate Molecular Mechanisms Involved in Host Plant Responses to Plasmodiophora brassicae |
title_sort | application of the nanostring ncounter system as an alternative method to investigate molecular mechanisms involved in host plant responses to plasmodiophora brassicae |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9779335/ https://www.ncbi.nlm.nih.gov/pubmed/36555223 http://dx.doi.org/10.3390/ijms232415581 |
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