Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus

Abstract Choroid plexus (ChP), a vascularized secretory epithelium located in all brain ventricles, plays critical roles in development, homeostasis and brain repair. Reverse transcription quantitative real-time PCR (RT-qPCR) is a popular and useful technique for measuring gene expression changes an...

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Autores principales:	Kim Hoa Ho, Annarita Patrizi
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/0c88f79c41da44129611577f3d79ab3e
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spelling	oai:doaj.org-article:0c88f79c41da44129611577f3d79ab3e2021-12-02T13:30:27ZAssessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus10.1038/s41598-021-82800-52045-2322https://doaj.org/article/0c88f79c41da44129611577f3d79ab3e2021-02-01T00:00:00Zhttps://doi.org/10.1038/s41598-021-82800-5https://doaj.org/toc/2045-2322Abstract Choroid plexus (ChP), a vascularized secretory epithelium located in all brain ventricles, plays critical roles in development, homeostasis and brain repair. Reverse transcription quantitative real-time PCR (RT-qPCR) is a popular and useful technique for measuring gene expression changes and also widely used in ChP studies. However, the reliability of RT-qPCR data is strongly dependent on the choice of reference genes, which are supposed to be stable across all samples. In this study, we validated the expression of 12 well established housekeeping genes in ChP in 2 independent experimental paradigms by using popular stability testing algorithms: BestKeeper, DeltaCq, geNorm and NormFinder. Rer1 and Rpl13a were identified as the most stable genes throughout mouse ChP development, while Hprt1 and Rpl27 were the most stable genes across conditions in a mouse sensory deprivation experiment. In addition, Rpl13a, Rpl27 and Tbp were mutually among the top five most stable genes in both experiments. Normalisation of Ttr and Otx2 expression levels using different housekeeping gene combinations demonstrated the profound effect of reference gene choice on target gene expression. Our study emphasized the importance of validating and selecting stable housekeeping genes under specific experimental conditions.Kim Hoa HoAnnarita PatriziNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 11, Iss 1, Pp 1-15 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Kim Hoa Ho Annarita Patrizi Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
description	Abstract Choroid plexus (ChP), a vascularized secretory epithelium located in all brain ventricles, plays critical roles in development, homeostasis and brain repair. Reverse transcription quantitative real-time PCR (RT-qPCR) is a popular and useful technique for measuring gene expression changes and also widely used in ChP studies. However, the reliability of RT-qPCR data is strongly dependent on the choice of reference genes, which are supposed to be stable across all samples. In this study, we validated the expression of 12 well established housekeeping genes in ChP in 2 independent experimental paradigms by using popular stability testing algorithms: BestKeeper, DeltaCq, geNorm and NormFinder. Rer1 and Rpl13a were identified as the most stable genes throughout mouse ChP development, while Hprt1 and Rpl27 were the most stable genes across conditions in a mouse sensory deprivation experiment. In addition, Rpl13a, Rpl27 and Tbp were mutually among the top five most stable genes in both experiments. Normalisation of Ttr and Otx2 expression levels using different housekeeping gene combinations demonstrated the profound effect of reference gene choice on target gene expression. Our study emphasized the importance of validating and selecting stable housekeeping genes under specific experimental conditions.
format	article
author	Kim Hoa Ho Annarita Patrizi
author_facet	Kim Hoa Ho Annarita Patrizi
author_sort	Kim Hoa Ho
title	Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
title_short	Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
title_full	Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
title_fullStr	Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
title_full_unstemmed	Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus
title_sort	assessment of common housekeeping genes as reference for gene expression studies using rt-qpcr in mouse choroid plexus
publisher	Nature Portfolio
publishDate	2021
url	https://doaj.org/article/0c88f79c41da44129611577f3d79ab3e
work_keys_str_mv	AT kimhoaho assessmentofcommonhousekeepinggenesasreferenceforgeneexpressionstudiesusingrtqpcrinmousechoroidplexus AT annaritapatrizi assessmentofcommonhousekeepinggenesasreferenceforgeneexpressionstudiesusingrtqpcrinmousechoroidplexus
_version_	1718392896927825920

Assessment of common housekeeping genes as reference for gene expression studies using RT-qPCR in mouse choroid plexus

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