Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability
Radish (Raphanus sativus) is an important cruciferous root crop with a close relationship to Chinese cabbage (Brassica rapa). RT-qPCR is used extensively to evaluate the expression levels of target genes, and accurate measurement of target gene expression with this method is determined by the valid...
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Frontiers Media S.A.
2017-09-01
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Series: | Frontiers in Plant Science |
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Online Access: | http://journal.frontiersin.org/article/10.3389/fpls.2017.01605/full |
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DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Mengmeng Duan Mengmeng Duan Jinglei Wang Xiaohui Zhang Haohui Yang Haiping Wang Yang Qiu Jiangping Song Yangdong Guo Xixiang Li |
spellingShingle |
Mengmeng Duan Mengmeng Duan Jinglei Wang Xiaohui Zhang Haohui Yang Haiping Wang Yang Qiu Jiangping Song Yangdong Guo Xixiang Li Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability Frontiers in Plant Science radish Chinese cabbage distant hybrid organs stress pistil development |
author_facet |
Mengmeng Duan Mengmeng Duan Jinglei Wang Xiaohui Zhang Haohui Yang Haiping Wang Yang Qiu Jiangping Song Yangdong Guo Xixiang Li |
author_sort |
Mengmeng Duan |
title |
Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability |
title_short |
Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability |
title_full |
Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability |
title_fullStr |
Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability |
title_full_unstemmed |
Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression Stability |
title_sort |
identification of optimal reference genes for expression analysis in radish (raphanus sativus l.) and its relatives based on expression stability |
publisher |
Frontiers Media S.A. |
series |
Frontiers in Plant Science |
issn |
1664-462X |
publishDate |
2017-09-01 |
description |
Radish (Raphanus sativus) is an important cruciferous root crop with a close relationship to Chinese cabbage (Brassica rapa). RT-qPCR is used extensively to evaluate the expression levels of target genes, and accurate measurement of target gene expression with this method is determined by the valid reference genes used for data nomalization in different experimental conditions. Screening for appropriate reference genes with stable expression based on RT-qPCR data is important for gene expression and functional analysis research in radish and its relatives. However, many researches have thought that almost no single reference gene is widely suitable for all experimental conditions, and few researchers have paid attention to the validation of reference genes in radish gene expression analysis. In the present study, 12 candidate reference genes were selected for analysis. Their expression in 28 samples, including 20 radish samples from different organs and conditions, four Chinese cabbage organs and four organs of their distant hybrid, was assessed by RT-qPCR and then five software tools—ΔCt, geNorm, NormFinder, BestKeeper and RefFinder—were used to compare their expression stability. The results showed that the most suitable reference genes were different in different organs and conditions. GAPDH, DSS1, and UP2 were optimal reference genes for gene expression analysis in all organs and conditions in radish. UPR, GSNOR1, and ACTIN2/7 were the most stable reference genes in different radish organs. UP2 and GAPDH were suitable reference genes for radish pistil development studies. RPII, UBC9, and GAPDH had the most stable expression in radish under various stresses. DSS1, UP2, and TEF2 were the optimal reference genes for Chinese cabbage organs, whereas TUA was optimal for the distant hybrid. UP2, and TEF2 were appropriate reference genes for all of the samples together. The optimal reference genes we identified, UP2, GAPDH, UPR, and GSNOR1 were verified by normalizing the expression patterns of YAB3, RPL, and FUL. These results will provide important information for selecting target reference genes in different research contexts and improve the accuracy and precision of gene expression analysis for radish, Chinese cabbage and their distant hybrid. |
topic |
radish Chinese cabbage distant hybrid organs stress pistil development |
url |
http://journal.frontiersin.org/article/10.3389/fpls.2017.01605/full |
work_keys_str_mv |
AT mengmengduan identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT mengmengduan identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT jingleiwang identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT xiaohuizhang identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT haohuiyang identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT haipingwang identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT yangqiu identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT jiangpingsong identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT yangdongguo identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability AT xixiangli identificationofoptimalreferencegenesforexpressionanalysisinradishraphanussativuslanditsrelativesbasedonexpressionstability |
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doaj-bfa363da2d9f4e23805064bd85db76632020-11-24T23:13:28ZengFrontiers Media S.A.Frontiers in Plant Science1664-462X2017-09-01810.3389/fpls.2017.01605266141Identification of Optimal Reference Genes for Expression Analysis in Radish (Raphanus sativus L.) and Its Relatives Based on Expression StabilityMengmeng Duan0Mengmeng Duan1Jinglei Wang2Xiaohui Zhang3Haohui Yang4Haiping Wang5Yang Qiu6Jiangping Song7Yangdong Guo8Xixiang Li9Key Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaDepartment of Vegetable Sciences, College of Horticulture, China Agricultural UniversityBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaDepartment of Vegetable Sciences, College of Horticulture, China Agricultural UniversityBeijing, ChinaKey Laboratory of Biology and Genetic Improvement of Horticultural Crops, Ministry of Agriculture, Institute of Vegetables and Flowers, Chinese Academy of Agricultural SciencesBeijing, ChinaRadish (Raphanus sativus) is an important cruciferous root crop with a close relationship to Chinese cabbage (Brassica rapa). RT-qPCR is used extensively to evaluate the expression levels of target genes, and accurate measurement of target gene expression with this method is determined by the valid reference genes used for data nomalization in different experimental conditions. Screening for appropriate reference genes with stable expression based on RT-qPCR data is important for gene expression and functional analysis research in radish and its relatives. However, many researches have thought that almost no single reference gene is widely suitable for all experimental conditions, and few researchers have paid attention to the validation of reference genes in radish gene expression analysis. In the present study, 12 candidate reference genes were selected for analysis. Their expression in 28 samples, including 20 radish samples from different organs and conditions, four Chinese cabbage organs and four organs of their distant hybrid, was assessed by RT-qPCR and then five software tools—ΔCt, geNorm, NormFinder, BestKeeper and RefFinder—were used to compare their expression stability. The results showed that the most suitable reference genes were different in different organs and conditions. GAPDH, DSS1, and UP2 were optimal reference genes for gene expression analysis in all organs and conditions in radish. UPR, GSNOR1, and ACTIN2/7 were the most stable reference genes in different radish organs. UP2 and GAPDH were suitable reference genes for radish pistil development studies. RPII, UBC9, and GAPDH had the most stable expression in radish under various stresses. DSS1, UP2, and TEF2 were the optimal reference genes for Chinese cabbage organs, whereas TUA was optimal for the distant hybrid. UP2, and TEF2 were appropriate reference genes for all of the samples together. The optimal reference genes we identified, UP2, GAPDH, UPR, and GSNOR1 were verified by normalizing the expression patterns of YAB3, RPL, and FUL. These results will provide important information for selecting target reference genes in different research contexts and improve the accuracy and precision of gene expression analysis for radish, Chinese cabbage and their distant hybrid.http://journal.frontiersin.org/article/10.3389/fpls.2017.01605/fullradishChinese cabbagedistant hybridorgansstresspistil development |