title: A tissue-based approach to selection of reference genes for quantitative real-time PCR in a sheep osteoporosis model
creator: Schulze, Felix
creator: Malhan, Deeksha
creator: EL Khassawna, Thaqif
creator: Heiss, Christian
creator: Seckinger, Anja
creator: Hose, Dirk
creator: Rösen-Wolff, Angela
subject: ddc-610
subject: 610 Medical sciences Medicine
description: Background: In order to better understand the multifactorial nature of osteoporosis, animal models are utilized and compared to healthy controls. Female sheep are well established as a model for osteoporosis induced by ovariectomy, calcium and vitamin D low diet, application of steroids, or a combination of these treatments. Transcriptional studies can be performed by applying quantitative real time PCR (RT-qPCR). RT-qPCR estimates mRNA-levels of target genes in relation to reference genes. A chosen set of reference genes should not show variation under experimental conditions. Currently, no standard reference genes are accepted for all tissue types and experimental conditions. Studies examining reference genes for sheep are rare and only one study described stable reference in mandibular bone. However, this type of bone differs from trabecular bone where most osteoporotic fractures occur. The present study aimed at identifying a set of reference genes for relative quantification of transcriptional activity of ovine spine bone and ovine in vitro differentiated mesenchymal stromal cells (MSC) for reliable comparability.    Methods: Twelve candidate reference genes belonging to different functional classes were selected and their expression was measured from cultured ovMSCs (n = 18) and ovine bone samples (n = 16), respectively. RefFinder was used to rank the candidate genes.    Results: We identified B2M, GAPDH, RPL19 and YWHAZ as the best combination of reference genes for normalization of RT-qPCR results for transcriptional analyses of these ovine samples.    Conclusion: This study demonstrates the importance of applying a set of reference genes for RT-qPCR analysis in sheep. Based on our data we recommend using four identified reference genes for relative quantification of gene expression studies in ovine bone or for in vitro experiments with osteogenically differentiated ovine MSCs.
publisher: BioMed Central ; Springer
date: 2017
type: Article
type: info:eu-repo/semantics/article
type: NonPeerReviewed
format: application/pdf
identifier: https://archiv.ub.uni-heidelberg.de/volltextserverhttps://archiv.ub.uni-heidelberg.de/volltextserver/24269/1/12864_2017_Article_4356.pdf
identifier: DOI:
identifier: urn:nbn:de:bsz:16-heidok-242697
identifier:   Schulze, Felix ; Malhan, Deeksha ; EL Khassawna, Thaqif ; Heiss, Christian ; Seckinger, Anja ; Hose, Dirk ; Rösen-Wolff, Angela  (2017) A tissue-based approach to selection of reference genes for quantitative real-time PCR in a sheep osteoporosis model.  BMC Genomics, 18 (975).  pp. 2-9.  ISSN 1471-2164     
relation: https://archiv.ub.uni-heidelberg.de/volltextserver/24269/
rights: info:eu-repo/semantics/openAccess
rights: Please see front page of the work (Sorry, Dublin Core plugin does not recognise license id)
language: eng