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- W4210363809 abstract "To underline the necessity of adequate reference genes for real-time quantitative polymerase chain reaction (RT-qPCR) and evaluate a novel tool for condition-specific reference gene selection.RT-qPCR is a commonly used experimental technique that allows for highly sensitive analysis of gene transcription. Moreover, the use of internal reference genes as a means for relative quantification has rendered RT-qPCR a straightforward method for a variety of sciences, including dentistry. However, the expressional stability of internal reference genes must be evaluated for every assay in order to account for possible quantification bias.Herein, we used the software tool RefGenes to identify putatively stable reference genes with the help of microarray datasets and evaluated them. Additionally, we propose an evidence-based workflow for adequate normalization of thusly identified genes. Human gingival fibroblasts (HGF-hTert), human acute leukemia-derived monocytes (THP-1), and telomerase immortalized gingival keratinocytes (TIGKs) were subjected to set-ups simulating various glycemic conditions and lipopolysaccharide challenges. Five common housekeeping genes (HKGs) and five genes from RefGenes were selected as targets and RT-qPCR was performed subsequently. Then, normalization algorithms Bestkeeper, Normfinder, and geNorm were used for further analysis of the putative reference gene stability.RefGenes-derived targets exhibited the highest stability values in THP-1 and TIGK cell lines. Moreover, unacceptable standard variations were observed for some common HKG like β-actin. However, common HKG exhibited good stability values in HGF-hTert cells.The results indicate that microarray-based preselection of putative reference genes is a valuable refinement for RT-qPCR studies. Accordingly, the present study proposes a straightforward workflow for evidence-based preselection and validation of internal reference genes." @default.
- W4210363809 created "2022-02-08" @default.
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- W4210363809 date "2022-02-01" @default.
- W4210363809 modified "2023-10-16" @default.
- W4210363809 title "Evidence‐based selection of reference genes for RT‐qPCR assays in periodontal research" @default.
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- W4210363809 doi "https://doi.org/10.1002/cre2.525" @default.
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