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Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR
In this study we evaluated eleven candidate reference genes in Avena sativa during compatible and incompatible interactions with two different pathotypes of Puccinia coronata f. sp. avenae in six time points post-inoculation. The identification of genes with high expression stability was performed b...
Autores principales: | , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Nature Publishing Group UK
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9418433/ https://www.ncbi.nlm.nih.gov/pubmed/36028746 http://dx.doi.org/10.1038/s41598-022-18746-z |
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author | Sowa, Sylwia Sozoniuk, Magdalena Toporowska, Joanna Kowalczyk, Krzysztof Paczos-Grzęda, Edyta |
author_facet | Sowa, Sylwia Sozoniuk, Magdalena Toporowska, Joanna Kowalczyk, Krzysztof Paczos-Grzęda, Edyta |
author_sort | Sowa, Sylwia |
collection | PubMed |
description | In this study we evaluated eleven candidate reference genes in Avena sativa during compatible and incompatible interactions with two different pathotypes of Puccinia coronata f. sp. avenae in six time points post-inoculation. The identification of genes with high expression stability was performed by four algorithms (geNorm, NormFinder, BestKeeper and ΔCt method). The results obtained confirmed that the combination of two genes would be sufficient for reliable normalization of the expression data. In general, the most stable in the tested plant-pathogen system were HNR (heterogeneous nuclear ribonucleoprotein 27C) and EF1A (elongation factor 1-alpha). ARF (ADP-ribosylation factor) and EIF4A (eukaryotic initiation factor 4A-3) could also be considered as exhibiting high expression stability. CYP (cyclophilin) was shown by all assessment methods to be the worst candidate for normalization in this dataset. To date, this is the first report of reference genes selection in A. sativa–P. coronata interaction system. Identified reference genes enable reliable and comprehensive RT-qPCR analysis of oat gene expression in response to crown rust infection. Understanding the molecular mechanisms involved in the host–pathogen interactions may expand knowledge of durable resistance strategies beneficial to modern oat breeding. |
format | Online Article Text |
id | pubmed-9418433 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Nature Publishing Group UK |
record_format | MEDLINE/PubMed |
spelling | pubmed-94184332022-08-28 Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR Sowa, Sylwia Sozoniuk, Magdalena Toporowska, Joanna Kowalczyk, Krzysztof Paczos-Grzęda, Edyta Sci Rep Article In this study we evaluated eleven candidate reference genes in Avena sativa during compatible and incompatible interactions with two different pathotypes of Puccinia coronata f. sp. avenae in six time points post-inoculation. The identification of genes with high expression stability was performed by four algorithms (geNorm, NormFinder, BestKeeper and ΔCt method). The results obtained confirmed that the combination of two genes would be sufficient for reliable normalization of the expression data. In general, the most stable in the tested plant-pathogen system were HNR (heterogeneous nuclear ribonucleoprotein 27C) and EF1A (elongation factor 1-alpha). ARF (ADP-ribosylation factor) and EIF4A (eukaryotic initiation factor 4A-3) could also be considered as exhibiting high expression stability. CYP (cyclophilin) was shown by all assessment methods to be the worst candidate for normalization in this dataset. To date, this is the first report of reference genes selection in A. sativa–P. coronata interaction system. Identified reference genes enable reliable and comprehensive RT-qPCR analysis of oat gene expression in response to crown rust infection. Understanding the molecular mechanisms involved in the host–pathogen interactions may expand knowledge of durable resistance strategies beneficial to modern oat breeding. Nature Publishing Group UK 2022-08-26 /pmc/articles/PMC9418433/ /pubmed/36028746 http://dx.doi.org/10.1038/s41598-022-18746-z Text en © The Author(s) 2022 https://creativecommons.org/licenses/by/4.0/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/ (https://creativecommons.org/licenses/by/4.0/) . |
spellingShingle | Article Sowa, Sylwia Sozoniuk, Magdalena Toporowska, Joanna Kowalczyk, Krzysztof Paczos-Grzęda, Edyta Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR |
title | Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR |
title_full | Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR |
title_fullStr | Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR |
title_full_unstemmed | Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR |
title_short | Validation of reference genes as an internal control for studying Avena sativa–Puccinia coronata interaction by RT-qPCR |
title_sort | validation of reference genes as an internal control for studying avena sativa–puccinia coronata interaction by rt-qpcr |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9418433/ https://www.ncbi.nlm.nih.gov/pubmed/36028746 http://dx.doi.org/10.1038/s41598-022-18746-z |
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