Validation of suitable genes for normalization of diurnal gene expression studies in Chenopodium quinoa

Nathaly Maldonado-Taipe; Dilan S. R. Patirange; Sandra M. Schmöckel; Christian Jung; Nazgol Emrani

doi:10.1371/journal.pone.0233821

Validation of suitable genes for normalization of diurnal gene expression studies in Chenopodium quinoa

Nathaly Maldonado-Taipe, Dilan S. R. Patirange, Sandra M. Schmöckel, Christian Jung, Nazgol Emrani

King Abdullah University of Science and Technology

Research output: Contribution to journal › Article › peer-review

8 Scopus citations

Abstract

Quinoa depicts high nutritional quality and abiotic stress resistance, attracting strong interest in the last years. To unravel the function of candidate genes for agronomically relevant traits, studying their transcriptional activities by RT-qPCR is an important experimental approach. The accuracy of such experiments strongly depends on precise data normalization. To date, validation of potential candidate genes for normalization of diurnal expression studies has not been performed in C. quinoa. We selected eight candidate genes based on transcriptome data and literature survey, including conventionally used reference genes. We used three statistical algorithms (BestKeeper, geNorm and NormFinder) to test their stability and added further validation by a simulation-based strategy. We demonstrated that using different reference genes, including those top ranked by stability, causes significant differences among the resulting diurnal expression patterns. Our results show that isocitrate dehydrogenase enzyme (IDH-A) and polypyrimidine tract-binding protein (PTB) are suitable genes to normalize diurnal expression data of two different quinoa accessions. Moreover, we validated our reference genes by normalizing two known diurnally regulated genes, BTC1 and BBX19. The validated reference genes obtained in this study will improve the accuracy of RT-qPCR data normalization and facilitate gene expression studies in quinoa.

Original language	English (US)
Pages (from-to)	e0233821
Journal	PLOS ONE
Volume	16
Issue number	3
DOIs	https://doi.org/10.1371/journal.pone.0233821
State	Published - Mar 11 2021

Bibliographical note

KAUST Repository Item: Exported on 2021-03-30
Acknowledged KAUST grant number(s): OSR-2016-CRG5-2966-02
Acknowledgements: This study was partially funded by the Competitive Research Grant of the King Abdullah University of Science and Technology, Saudi Arabia
awarded to CJ (Grant number: OSR-2016-CRG5-2966-02). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. There was no additional external funding received for this study. The rest of the costs were covered internally by the institute budget of the Plant Breeding Institute, Kiel University. We acknowledge the financial support by State of Schleswig-Holstein, Germany within the funding program “Open Access Publikationsfonds”.

ASJC Scopus subject areas

General Agricultural and Biological Sciences
General Biochemistry, Genetics and Molecular Biology
General Medicine

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10.1371/journal.pone.0233821

https://repository.kaust.edu.sa/bitstream/10754/667731/3/journal.pone.0233821%20%281%29.pdf

Cite this

@article{71c7ba09a4ab4ac08d6182aabde2488c,

title = "Validation of suitable genes for normalization of diurnal gene expression studies in Chenopodium quinoa",

abstract = "Quinoa depicts high nutritional quality and abiotic stress resistance, attracting strong interest in the last years. To unravel the function of candidate genes for agronomically relevant traits, studying their transcriptional activities by RT-qPCR is an important experimental approach. The accuracy of such experiments strongly depends on precise data normalization. To date, validation of potential candidate genes for normalization of diurnal expression studies has not been performed in C. quinoa. We selected eight candidate genes based on transcriptome data and literature survey, including conventionally used reference genes. We used three statistical algorithms (BestKeeper, geNorm and NormFinder) to test their stability and added further validation by a simulation-based strategy. We demonstrated that using different reference genes, including those top ranked by stability, causes significant differences among the resulting diurnal expression patterns. Our results show that isocitrate dehydrogenase enzyme (IDH-A) and polypyrimidine tract-binding protein (PTB) are suitable genes to normalize diurnal expression data of two different quinoa accessions. Moreover, we validated our reference genes by normalizing two known diurnally regulated genes, BTC1 and BBX19. The validated reference genes obtained in this study will improve the accuracy of RT-qPCR data normalization and facilitate gene expression studies in quinoa.",

author = "Nathaly Maldonado-Taipe and Patirange, {Dilan S. R.} and Schm{\"o}ckel, {Sandra M.} and Christian Jung and Nazgol Emrani",

note = "KAUST Repository Item: Exported on 2021-03-30 Acknowledged KAUST grant number(s): OSR-2016-CRG5-2966-02 Acknowledgements: This study was partially funded by the Competitive Research Grant of the King Abdullah University of Science and Technology, Saudi Arabia awarded to CJ (Grant number: OSR-2016-CRG5-2966-02). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. There was no additional external funding received for this study. The rest of the costs were covered internally by the institute budget of the Plant Breeding Institute, Kiel University. We acknowledge the financial support by State of Schleswig-Holstein, Germany within the funding program “Open Access Publikationsfonds”.",

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doi = "10.1371/journal.pone.0233821",

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T1 - Validation of suitable genes for normalization of diurnal gene expression studies in Chenopodium quinoa

AU - Maldonado-Taipe, Nathaly

AU - Patirange, Dilan S. R.

AU - Schmöckel, Sandra M.

AU - Jung, Christian

AU - Emrani, Nazgol

N1 - KAUST Repository Item: Exported on 2021-03-30 Acknowledged KAUST grant number(s): OSR-2016-CRG5-2966-02 Acknowledgements: This study was partially funded by the Competitive Research Grant of the King Abdullah University of Science and Technology, Saudi Arabia awarded to CJ (Grant number: OSR-2016-CRG5-2966-02). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. There was no additional external funding received for this study. The rest of the costs were covered internally by the institute budget of the Plant Breeding Institute, Kiel University. We acknowledge the financial support by State of Schleswig-Holstein, Germany within the funding program “Open Access Publikationsfonds”.

PY - 2021/3/11

Y1 - 2021/3/11

N2 - Quinoa depicts high nutritional quality and abiotic stress resistance, attracting strong interest in the last years. To unravel the function of candidate genes for agronomically relevant traits, studying their transcriptional activities by RT-qPCR is an important experimental approach. The accuracy of such experiments strongly depends on precise data normalization. To date, validation of potential candidate genes for normalization of diurnal expression studies has not been performed in C. quinoa. We selected eight candidate genes based on transcriptome data and literature survey, including conventionally used reference genes. We used three statistical algorithms (BestKeeper, geNorm and NormFinder) to test their stability and added further validation by a simulation-based strategy. We demonstrated that using different reference genes, including those top ranked by stability, causes significant differences among the resulting diurnal expression patterns. Our results show that isocitrate dehydrogenase enzyme (IDH-A) and polypyrimidine tract-binding protein (PTB) are suitable genes to normalize diurnal expression data of two different quinoa accessions. Moreover, we validated our reference genes by normalizing two known diurnally regulated genes, BTC1 and BBX19. The validated reference genes obtained in this study will improve the accuracy of RT-qPCR data normalization and facilitate gene expression studies in quinoa.

AB - Quinoa depicts high nutritional quality and abiotic stress resistance, attracting strong interest in the last years. To unravel the function of candidate genes for agronomically relevant traits, studying their transcriptional activities by RT-qPCR is an important experimental approach. The accuracy of such experiments strongly depends on precise data normalization. To date, validation of potential candidate genes for normalization of diurnal expression studies has not been performed in C. quinoa. We selected eight candidate genes based on transcriptome data and literature survey, including conventionally used reference genes. We used three statistical algorithms (BestKeeper, geNorm and NormFinder) to test their stability and added further validation by a simulation-based strategy. We demonstrated that using different reference genes, including those top ranked by stability, causes significant differences among the resulting diurnal expression patterns. Our results show that isocitrate dehydrogenase enzyme (IDH-A) and polypyrimidine tract-binding protein (PTB) are suitable genes to normalize diurnal expression data of two different quinoa accessions. Moreover, we validated our reference genes by normalizing two known diurnally regulated genes, BTC1 and BBX19. The validated reference genes obtained in this study will improve the accuracy of RT-qPCR data normalization and facilitate gene expression studies in quinoa.

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JO - PLOS ONE

JF - PLOS ONE

IS - 3

ER -

Validation of suitable genes for normalization of diurnal gene expression studies in Chenopodium quinoa

Abstract

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ASJC Scopus subject areas

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