ArticleEthiopian journal of health sciences2018
Normalization of Gene Expression by Quantitative RT-PCR in Human Cell Line: comparison of 12 Endogenous Reference Genes.
Article in Ethiopian journal of health sciences, 2018. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 23 papers.
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Who cites it
23 citing papers in PubMed, 32 citations in OpenAlex.
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- Identification of optimal reference genes in golden Syrian hamster with ethanol- and palmitoleic acid-induced acute pancreatitis using quantitative real-time polymerase chain reaction.Animal models and experimental medicine · 2023Article
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- A Strategy for the Selection of RT-qPCR Reference Genes Based on Publicly Available Transcriptomic Datasets.Biomedicines · 2023Article
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- Field Evaluation of a Duplex RT-RAA Assay for Rapid Detection of SARS-CoV-2 - Hebei Province, China, January 2021.China CDC weekly · 2022Article
- Screening and stability analysis of reference genes in fasting caecotrophy model in rabbits.Molecular biology reports · 2022Article
- NR4A1 inhibits the epithelial-mesenchymal transition of hepatic stellate cells: Involvement of TGF-β-Smad2/3/4-ZEB signaling.Open life sciences · 2022Article
- Identification of Reference Genes for Expression Studies in the Whole-Blood from Three Cattle Breeds under Two States of Livestock Weather Safety.Animals : an open access journal from MDPI · 2021Article
- Adverse Effect of Circadian Rhythm Disorder on Reparative Angiogenesis in Hind Limb Ischemia.Journal of the American Heart Association · 2021Article
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- From Donor to the Lab: A Fascinating Journey of Primary Cell Lines.Frontiers in cell and developmental biology · 2021Review
Corrections and comments
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Authors and funding
1 author at 1 institution in 2 countries.
Funding
No grant is acknowledged in the PubMed record.
Abstract
backgroundPolymerase Chain Reaction (PCR) has become an important diagnostic and research tool of modern molecular biology globally. Real-time PCR allows for rapid and reliable quantification of mRNA transcription. Reference genes are used as internal reaction control to normalise mRNA levels between different samples in order to allow for an exact comparison of mRNA transcription level.
methodsIn this study, twelve commonly used human reference genes were investigated in Human Embryonic Kidney Cell Lines (HEK293) using real-time qPCR with SYBR green. The genes included beta-2-microglobulin (B2M), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), succinate dehydrogenase complex subunit A (SDHA), and tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein zeta polypeptide (YWHAZ). The stability of these reference genes was investigated using the geNorm application.
resultsThe range of expression stability in the genes analysed was (from the most stable to the least stable): UBC, TOP1, ATP5B, CYC1, GAPDH, SDHA, YWHAZ, CTB, 18S, EIFA-2, B2M and RPL13A. The optimal number of reference targets in the experiment was calculated to be 2 (geNorm V<0.15) when comparing a normalization factor based on the 2 or 3 most stable targets).
conclusionThe expression stability varied greatly between the 12 candidate reference genes. UBC, TOP1, ATP5B, CYC1 and GAPDH respectively showed the highest stability in HEK293 cells based on both expression stability and expression level. Overall, our data suggest that UBC and TOP1show the least variation and the highest expression stability. This report validates the need for rational selection of reference genes for data normalization to ensure accuracy of quantitative PCR assays.
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