Forskning ved Københavns Universitet - Københavns Universitet

Forside

Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

Standard

Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro. / Hersom, Maria; Goldeman, Charlotte; Pretzer, Natasia; Brodin, Birger.

I: Molecular and Cellular Neuroscience, Bind 93, 12.2018, s. 27-35.

Publikation: Bidrag til tidsskriftTidsskriftartikelForskningfagfællebedømt

Harvard

Hersom, M, Goldeman, C, Pretzer, N & Brodin, B 2018, 'Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro', Molecular and Cellular Neuroscience, bind 93, s. 27-35. https://doi.org/10.1016/j.mcn.2018.10.001

APA

Hersom, M., Goldeman, C., Pretzer, N., & Brodin, B. (2018). Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro. Molecular and Cellular Neuroscience, 93, 27-35. https://doi.org/10.1016/j.mcn.2018.10.001

Vancouver

Hersom M, Goldeman C, Pretzer N, Brodin B. Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro. Molecular and Cellular Neuroscience. 2018 dec;93:27-35. https://doi.org/10.1016/j.mcn.2018.10.001

Author

Hersom, Maria ; Goldeman, Charlotte ; Pretzer, Natasia ; Brodin, Birger. / Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro. I: Molecular and Cellular Neuroscience. 2018 ; Bind 93. s. 27-35.

Bibtex

@article{c60821c152ad4d7fbf4131bd2534b31a,
title = "Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro",
abstract = "BACKGROUND: The genes encoding β-actin and GAPDH are two of the most commonly used reference genes for normalization in in vitro blood-brain barrier studies. Studies have, however, shown that these reference genes might not always be the best choice. The aim of the present study was to evaluate 10 reference genes for use in mRNA profiling studies in primary cultures of brain endothelial cells of bovine origin.METHODS: Gene evaluations were performed by qPCR in mono-culture and in co-cultures with astrocytes. The expression of reference genes was furthermore investigated during culture. Qbase+ software was used to analyze the stability of the tested genes and for determinations of the optimal number of reference genes.RESULTS: The stability of the reference genes varied between the culture configurations, but for all culture configurations we found that the optimal number of reference genes were two. PMM-1, RPL13A and β-actin were the most stable genes in mono-cultures, non-contact co-culture and contact co-culture respectively. For studies comparing gene expression between different culture configurations the optimal number of reference genes was three and RPL13A was found to be most stable. During cell culture a number of four reference genes were found to be optimal and YWHAZ was found to be the most stable gene. β-actin and GAPDH were found to be the least stable genes during culture.CONCLUSION: Overall we found that the validation of reference genes was important in order to normalize target gene expression correctly, and suggest sets of reference genes to be used under different experimental conditions, in order to quantify mRNA transcript levels in blood-brain barrier cell models correctly.",
author = "Maria Hersom and Charlotte Goldeman and Natasia Pretzer and Birger Brodin",
note = "Copyright {\textcopyright} 2018 Elsevier Inc. All rights reserved.",
year = "2018",
month = dec,
doi = "10.1016/j.mcn.2018.10.001",
language = "English",
volume = "93",
pages = "27--35",
journal = "Molecular and Cellular Neuroscience",
issn = "1044-7431",
publisher = "Academic Press",

}

RIS

TY - JOUR

T1 - Validation of reference genes for normalization of real-time quantitative PCR studies of gene expression in brain capillary endothelial cells cultured in vitro

AU - Hersom, Maria

AU - Goldeman, Charlotte

AU - Pretzer, Natasia

AU - Brodin, Birger

N1 - Copyright © 2018 Elsevier Inc. All rights reserved.

PY - 2018/12

Y1 - 2018/12

N2 - BACKGROUND: The genes encoding β-actin and GAPDH are two of the most commonly used reference genes for normalization in in vitro blood-brain barrier studies. Studies have, however, shown that these reference genes might not always be the best choice. The aim of the present study was to evaluate 10 reference genes for use in mRNA profiling studies in primary cultures of brain endothelial cells of bovine origin.METHODS: Gene evaluations were performed by qPCR in mono-culture and in co-cultures with astrocytes. The expression of reference genes was furthermore investigated during culture. Qbase+ software was used to analyze the stability of the tested genes and for determinations of the optimal number of reference genes.RESULTS: The stability of the reference genes varied between the culture configurations, but for all culture configurations we found that the optimal number of reference genes were two. PMM-1, RPL13A and β-actin were the most stable genes in mono-cultures, non-contact co-culture and contact co-culture respectively. For studies comparing gene expression between different culture configurations the optimal number of reference genes was three and RPL13A was found to be most stable. During cell culture a number of four reference genes were found to be optimal and YWHAZ was found to be the most stable gene. β-actin and GAPDH were found to be the least stable genes during culture.CONCLUSION: Overall we found that the validation of reference genes was important in order to normalize target gene expression correctly, and suggest sets of reference genes to be used under different experimental conditions, in order to quantify mRNA transcript levels in blood-brain barrier cell models correctly.

AB - BACKGROUND: The genes encoding β-actin and GAPDH are two of the most commonly used reference genes for normalization in in vitro blood-brain barrier studies. Studies have, however, shown that these reference genes might not always be the best choice. The aim of the present study was to evaluate 10 reference genes for use in mRNA profiling studies in primary cultures of brain endothelial cells of bovine origin.METHODS: Gene evaluations were performed by qPCR in mono-culture and in co-cultures with astrocytes. The expression of reference genes was furthermore investigated during culture. Qbase+ software was used to analyze the stability of the tested genes and for determinations of the optimal number of reference genes.RESULTS: The stability of the reference genes varied between the culture configurations, but for all culture configurations we found that the optimal number of reference genes were two. PMM-1, RPL13A and β-actin were the most stable genes in mono-cultures, non-contact co-culture and contact co-culture respectively. For studies comparing gene expression between different culture configurations the optimal number of reference genes was three and RPL13A was found to be most stable. During cell culture a number of four reference genes were found to be optimal and YWHAZ was found to be the most stable gene. β-actin and GAPDH were found to be the least stable genes during culture.CONCLUSION: Overall we found that the validation of reference genes was important in order to normalize target gene expression correctly, and suggest sets of reference genes to be used under different experimental conditions, in order to quantify mRNA transcript levels in blood-brain barrier cell models correctly.

U2 - 10.1016/j.mcn.2018.10.001

DO - 10.1016/j.mcn.2018.10.001

M3 - Journal article

C2 - 30315869

VL - 93

SP - 27

EP - 35

JO - Molecular and Cellular Neuroscience

JF - Molecular and Cellular Neuroscience

SN - 1044-7431

ER -

ID: 204042901