Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach
BACKGROUND: Quantitative PCR (qPCR) is a common method for quantifying mRNA expression. Given the heterogeneity present in tumor tissues, it is crucial to normalize target mRNA expression data using appropriate reference genes that are stably expressed under a variety of pathological and experimenta...
| Main Authors: | , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
BioMed Central
2017
|
| Online Access: | http://psasir.upm.edu.my/id/eprint/62931/1/Reference%20gene%20validation%20for%20gene%20expression%20normalization%20in%20canine%20osteosarcoma.pdf |
| _version_ | 1796977672195145728 |
|---|---|
| author | Selvarajah, Gayathri Thevi Bonestroo, Floor A. S. Timmermans Sprang, Elpetra P. M. Kirpensteijn, Jolle Mol, Jan A. |
| author_facet | Selvarajah, Gayathri Thevi Bonestroo, Floor A. S. Timmermans Sprang, Elpetra P. M. Kirpensteijn, Jolle Mol, Jan A. |
| author_sort | Selvarajah, Gayathri Thevi |
| collection | UPM |
| description | BACKGROUND: Quantitative PCR (qPCR) is a common method for quantifying mRNA expression. Given the heterogeneity present in tumor tissues, it is crucial to normalize target mRNA expression data using appropriate reference genes that are stably expressed under a variety of pathological and experimental conditions. No studies have validated specific reference genes in canine osteosarcoma (OS). Previous gene expression studies involving canine OS have used one or two reference genes to normalize gene expression. This study aimed to validate a panel of reference genes commonly used for normalization of canine OS gene expression data using the geNorm algorithm. qPCR analysis of nine canine reference genes was performed on 40 snap-frozen primary OS tumors and seven cell lines.
RESULTS: Tumors with a variety of clinical and pathological characteristics were selected. Gene expression stability and the optimal number of reference genes for gene expression normalization were calculated. RPS5 and HNRNPH were highly stable among OS cell lines, while RPS5 and RPS19 were the best combination for primary tumors. Pairwise variation analysis recommended four and two reference genes for optimal normalization of the expression data of canine OS tumors and cell lines, respectively.
CONCLUSIONS: Appropriate combinations of reference genes are recommended to normalize mRNA levels in canine OS tumors and cell lines to facilitate standardized and reliable quantification of target gene expression, which is essential for investigating key genes involved in canine OS metastasis and for comparative biomarker discovery. |
| first_indexed | 2024-03-06T09:43:31Z |
| format | Article |
| id | upm.eprints-62931 |
| institution | Universiti Putra Malaysia |
| language | English |
| last_indexed | 2024-03-06T09:43:31Z |
| publishDate | 2017 |
| publisher | BioMed Central |
| record_format | dspace |
| spelling | upm.eprints-629312018-09-28T10:37:22Z http://psasir.upm.edu.my/id/eprint/62931/ Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach Selvarajah, Gayathri Thevi Bonestroo, Floor A. S. Timmermans Sprang, Elpetra P. M. Kirpensteijn, Jolle Mol, Jan A. BACKGROUND: Quantitative PCR (qPCR) is a common method for quantifying mRNA expression. Given the heterogeneity present in tumor tissues, it is crucial to normalize target mRNA expression data using appropriate reference genes that are stably expressed under a variety of pathological and experimental conditions. No studies have validated specific reference genes in canine osteosarcoma (OS). Previous gene expression studies involving canine OS have used one or two reference genes to normalize gene expression. This study aimed to validate a panel of reference genes commonly used for normalization of canine OS gene expression data using the geNorm algorithm. qPCR analysis of nine canine reference genes was performed on 40 snap-frozen primary OS tumors and seven cell lines. RESULTS: Tumors with a variety of clinical and pathological characteristics were selected. Gene expression stability and the optimal number of reference genes for gene expression normalization were calculated. RPS5 and HNRNPH were highly stable among OS cell lines, while RPS5 and RPS19 were the best combination for primary tumors. Pairwise variation analysis recommended four and two reference genes for optimal normalization of the expression data of canine OS tumors and cell lines, respectively. CONCLUSIONS: Appropriate combinations of reference genes are recommended to normalize mRNA levels in canine OS tumors and cell lines to facilitate standardized and reliable quantification of target gene expression, which is essential for investigating key genes involved in canine OS metastasis and for comparative biomarker discovery. BioMed Central 2017-11-25 Article PeerReviewed text en http://psasir.upm.edu.my/id/eprint/62931/1/Reference%20gene%20validation%20for%20gene%20expression%20normalization%20in%20canine%20osteosarcoma.pdf Selvarajah, Gayathri Thevi and Bonestroo, Floor A. S. and Timmermans Sprang, Elpetra P. M. and Kirpensteijn, Jolle and Mol, Jan A. (2017) Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach. BMC Veterinary Research, 13 (354). pp. 1-8. ISSN 1746-6148 https://bmcvetres.biomedcentral.com/track/pdf/10.1186/s12917-017-1281-3 10.1186/s12917-017-1281-3 |
| spellingShingle | Selvarajah, Gayathri Thevi Bonestroo, Floor A. S. Timmermans Sprang, Elpetra P. M. Kirpensteijn, Jolle Mol, Jan A. Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach |
| title | Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach |
| title_full | Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach |
| title_fullStr | Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach |
| title_full_unstemmed | Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach |
| title_short | Reference gene validation for gene expression normalization in canine osteosarcoma: a geNorm algorithm approach |
| title_sort | reference gene validation for gene expression normalization in canine osteosarcoma a genorm algorithm approach |
| url | http://psasir.upm.edu.my/id/eprint/62931/1/Reference%20gene%20validation%20for%20gene%20expression%20normalization%20in%20canine%20osteosarcoma.pdf |
| work_keys_str_mv | AT selvarajahgayathrithevi referencegenevalidationforgeneexpressionnormalizationincanineosteosarcomaagenormalgorithmapproach AT bonestrooflooras referencegenevalidationforgeneexpressionnormalizationincanineosteosarcomaagenormalgorithmapproach AT timmermanssprangelpetrapm referencegenevalidationforgeneexpressionnormalizationincanineosteosarcomaagenormalgorithmapproach AT kirpensteijnjolle referencegenevalidationforgeneexpressionnormalizationincanineosteosarcomaagenormalgorithmapproach AT moljana referencegenevalidationforgeneexpressionnormalizationincanineosteosarcomaagenormalgorithmapproach |