A systemic approach to screening high-throughput RT-qPCR data for a suitable set of reference circulating miRNAs
Background The consensus on how to choose a reference gene for serum or plasma miRNA expression qPCR studies has not been reached and none of the potential candidates have yet been convincingly validated. We proposed a new in silico approach of finding a suitable reference for human, circulating miRNAs and identified a new set of endogenous reference miRNA based on miRNA profiling experiments from Gene Expression Omnibus. We used 3 known normalization algorithms (NormFinder, BestKeeper, GeNorm) to calculate a new normalization score. We searched for a universal set of endogenous miRNAs and validated our findings on 2 new datasets using our approach. Results We discovered and validated a set of 13 miRNAs (miR-222, miR-92a, miR-27a, miR-17, miR-24, miR-320a, miR-25, miR-126, miR-19b, miR-199a-3p, miR-30b, miR-30c, miR-374a) that can be used to create a reliable reference combination of 3 miRNAs. We showed that on average the mean of 3 miRNAs (p = 0.0002) and 2 miRNAs (p = 0.0031) were a better reference than single miRNA. The arithmetic means of 3 miRNAs: miR-24, miR-222 and miR-27a was shown to be the most stable combination of 3 miRNAs in validation sets. Conclusions No single miRNA was suitable as a universal reference in serum miRNA qPCR profiling, but it was possible to designate a set of miRNAs, which consistently contributed to most stable combinations..
| Medienart: |
E-Artikel |
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| Erscheinungsjahr: |
2020 |
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| Erschienen: |
2020 |
| Enthalten in: |
Zur Gesamtaufnahme - volume:21 |
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| Enthalten in: |
BMC genomics - 21(2020), 1 vom: 31. Jan. |
| Sprache: |
Englisch |
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| Beteiligte Personen: |
Pagacz, Konrad [VerfasserIn] |
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| Links: |
Volltext [kostenfrei] |
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| Anmerkungen: |
© The Author(s). 2020 |
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| doi: |
10.1186/s12864-020-6530-3 |
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| funding: |
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| Förderinstitution / Projekttitel: |
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| 520 | |a Background The consensus on how to choose a reference gene for serum or plasma miRNA expression qPCR studies has not been reached and none of the potential candidates have yet been convincingly validated. We proposed a new in silico approach of finding a suitable reference for human, circulating miRNAs and identified a new set of endogenous reference miRNA based on miRNA profiling experiments from Gene Expression Omnibus. We used 3 known normalization algorithms (NormFinder, BestKeeper, GeNorm) to calculate a new normalization score. We searched for a universal set of endogenous miRNAs and validated our findings on 2 new datasets using our approach. Results We discovered and validated a set of 13 miRNAs (miR-222, miR-92a, miR-27a, miR-17, miR-24, miR-320a, miR-25, miR-126, miR-19b, miR-199a-3p, miR-30b, miR-30c, miR-374a) that can be used to create a reliable reference combination of 3 miRNAs. We showed that on average the mean of 3 miRNAs (p = 0.0002) and 2 miRNAs (p = 0.0031) were a better reference than single miRNA. The arithmetic means of 3 miRNAs: miR-24, miR-222 and miR-27a was shown to be the most stable combination of 3 miRNAs in validation sets. Conclusions No single miRNA was suitable as a universal reference in serum miRNA qPCR profiling, but it was possible to designate a set of miRNAs, which consistently contributed to most stable combinations. | ||
| 700 | 1 | |a Kucharski, Przemyslaw |4 aut | |
| 700 | 1 | |a Smyczynska, Urszula |4 aut | |
| 700 | 1 | |a Grabia, Szymon |4 aut | |
| 700 | 1 | |a Chowdhury, Dipanjan |4 aut | |
| 700 | 1 | |a Fendler, Wojciech |4 aut | |
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