ArticleBMC immunology2020
Reliable reference genes for the quantification of mRNA in human T-cells and PBMCs stimulated with live influenza virus.
Article in BMC immunology, 2020. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 24 papers.
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24 citing papers in PubMed, 34 citations in OpenAlex.
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- Identification of Optimal Reference Genes for qRT-PCR Normalization for Physical Activity Intervention and Omega-3 Fatty Acids Supplementation in Humans.International journal of molecular sciences · 2023Article
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- Transcriptomic profiles of multiple organ dysfunction syndrome phenotypes in pediatric critical influenza.Frontiers in immunology · 2023Article
- A quest for cytocompatible metal organic frameworks in non-viral gene therapy: Relevance of zeolitic imidazolate framework-8.Biomaterials and biosystems · 2022Article
- Dual roles of oxostephanine as an Aurora kinase inhibitor and angiogenesis suppressor.International journal of molecular medicine · 2022Article
- At Embryo Implantation Site IL-35 Secreted by Trophoblast, Polarizing T Cells towards IL-35+ IL-10+ IL-4+ Th2-Type Cells, Could Favour Fetal Allograft Tolerance and Pregnancy Success.International journal of molecular sciences · 2022Article
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundQuantitative PCR (qPCR) is a powerful tool that is particularly well-suited to measure mRNA levels in clinical samples, especially those with relatively low cell counts. However, a caveat of this approach is that reliable, stably expressed reference (housekeeping) genes are vital in order to ensure reproducibility and appropriate biological inference. In this study, we evaluated the expression stability of six reference genes in peripheral blood mononuclear cells (PBMCs) and isolated CD3
resultsReference gene expression varied significantly depending on cell type and stimulation conditions, but not age. Using the comparative ΔCt method, and the previously published software BestKeeper, NormFinder, and geNorm, we show that in PBMCs and T-cells, UBE2D2 and RPS18 were the most stable reference genes, followed by ACTB; however, the expression of UBE2D2 and RPS18 was found to increase with viral stimulation in isolated T-cells, while ACTB expression did not change significantly. No age-related differences in stability were observed for any gene
conclusionsThis study suggests the use of a combination of UBE2D2, RPS18, and ACTB for the study of influenza responses in PBMCs and T-cells, although ACTB alone may be the most optimal choice if choosing to compare target gene expression before and after viral stimulation. Both GAPDH and RPL13a were found to be poor reference genes and should be avoided for studies of this nature.
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