Comparison of methods for relative quantification of gene ex- pression using real-time PCR
Quantitative real-time PCR (qPCR) has become a widely used tool for quantifying gene expression. Several methods for relative quantification have been developed, enabling rapid and reliable detection and quantification of specific nucleic acids. These methods, based on qPCR include: the standard cur...
| Main Authors: | , , , |
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| Format: | Article |
| Language: | English |
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University of Ljubljana Press (Založba Univerze v Ljubljani)
2012-12-01
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| Series: | Acta Agriculturae Slovenica |
| Subjects: | |
| Online Access: | https://journals.uni-lj.si/aas/article/view/13706 |
| _version_ | 1826532305815994368 |
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| author | Luka BOLHA Daliborka DUŠANIĆ Mojca NARAT Irena OVEN |
| author_facet | Luka BOLHA Daliborka DUŠANIĆ Mojca NARAT Irena OVEN |
| author_sort | Luka BOLHA |
| collection | DOAJ |
| description | Quantitative real-time PCR (qPCR) has become a widely used tool for quantifying gene expression. Several methods for relative quantification have been developed, enabling rapid and reliable detection and quantification of specific nucleic acids. These methods, based on qPCR include: the standard curve method, the efficiency calibrated method and the 2−ΔΔCq method. Here we analyzed if these three methods generate comparable results. To evaluate their performance, we analyzed the expression of the nuclease gene MS53_0284 from Mycoplasma synoviae type strain WVU 1853 during in vitro infection of CEC-32 cells, using qPCR. As determined, all three methods generated comparable and reliable results when all necessary conditions were fulfiled. Also, the efficiency calibrated and the standard curve methods were more suitable for quantifying small differences in relative gene expression than the 2−ΔΔCq method. |
| first_indexed | 2024-03-08T23:24:00Z |
| format | Article |
| id | doaj.art-91fd8705724c408088ca0c083d68679f |
| institution | Directory Open Access Journal |
| issn | 1854-1941 |
| language | English |
| last_indexed | 2025-03-14T01:49:08Z |
| publishDate | 2012-12-01 |
| publisher | University of Ljubljana Press (Založba Univerze v Ljubljani) |
| record_format | Article |
| series | Acta Agriculturae Slovenica |
| spelling | doaj.art-91fd8705724c408088ca0c083d68679f2025-03-12T12:21:37ZengUniversity of Ljubljana Press (Založba Univerze v Ljubljani)Acta Agriculturae Slovenica1854-19412012-12-0110029710610.14720/aas.2012.100.2.1370620101Comparison of methods for relative quantification of gene ex- pression using real-time PCRLuka BOLHA0Daliborka DUŠANIĆ1Mojca NARAT2Irena OVEN3Univ. of Ljubljana, Biotechnical Fac., Dept. of Animal Science, Groblje 3, SI-1230 Domžale, SloveniaUniv. of Ljubljana, Biotechnical Fac., Dept. of Animal Science, Groblje 3, SI-1230 Domžale, SloveniaUniv. of Ljubljana, Biotechnical Fac., Dept. of Animal Science, Groblje 3, SI-1230 Domžale, SloveniaUniv. of Ljubljana, Biotechnical Fac., Dept. of Animal Science, Groblje 3, SI-1230 Domžale, SloveniaQuantitative real-time PCR (qPCR) has become a widely used tool for quantifying gene expression. Several methods for relative quantification have been developed, enabling rapid and reliable detection and quantification of specific nucleic acids. These methods, based on qPCR include: the standard curve method, the efficiency calibrated method and the 2−ΔΔCq method. Here we analyzed if these three methods generate comparable results. To evaluate their performance, we analyzed the expression of the nuclease gene MS53_0284 from Mycoplasma synoviae type strain WVU 1853 during in vitro infection of CEC-32 cells, using qPCR. As determined, all three methods generated comparable and reliable results when all necessary conditions were fulfiled. Also, the efficiency calibrated and the standard curve methods were more suitable for quantifying small differences in relative gene expression than the 2−ΔΔCq method.https://journals.uni-lj.si/aas/article/view/13706molecular geneticsgenesgene expressionquantitative real-time pcrmethods |
| spellingShingle | Luka BOLHA Daliborka DUŠANIĆ Mojca NARAT Irena OVEN Comparison of methods for relative quantification of gene ex- pression using real-time PCR Acta Agriculturae Slovenica molecular genetics genes gene expression quantitative real-time pcr methods |
| title | Comparison of methods for relative quantification of gene ex- pression using real-time PCR |
| title_full | Comparison of methods for relative quantification of gene ex- pression using real-time PCR |
| title_fullStr | Comparison of methods for relative quantification of gene ex- pression using real-time PCR |
| title_full_unstemmed | Comparison of methods for relative quantification of gene ex- pression using real-time PCR |
| title_short | Comparison of methods for relative quantification of gene ex- pression using real-time PCR |
| title_sort | comparison of methods for relative quantification of gene ex pression using real time pcr |
| topic | molecular genetics genes gene expression quantitative real-time pcr methods |
| url | https://journals.uni-lj.si/aas/article/view/13706 |
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