Evaluation of Reference Genes for Quantitative Real-time PCR in Wharton's Jelly-derived Mesenchymal Stem Cells After Lentiviral Transduction and Differentiation

Overview

Journal Mol Biol Rep

Specialty Molecular Biology

Date 2019 Nov 30

PMID 31781918

Citations 12

Authors

P Borkowska

A Zielinska

M Paul-Samojedny

R Stojko

J Kowalski

Affiliations

Soon will be listed here.

Abstract

Quantitative real time reverse transcription PCR, qRT-PCR, is one of the most important techniques for assessing the level of gene expression. Selecting the correct reference gene to normalize the results is a key step in this method. Inaccurate data can be generated if the correct reference gene is not selected. The level of the expression of reference genes is tissue-variable, and in the case of mesenchymal stem cells (MSC), it can be different depending on the source of their origin. The aim of this study was to select the reference gene for Wharton's Jelly-derived MSC (WJ- MSC) that were undergoing transduction and differentiation. In this work, the expression of 32 genes was analyzed, of which two (RPS17 and 18S rRNA), which had the most stable expression level, were selected. A comparative analysis of the expression stability of the selected genes was then performed with the genes that are most commonly used in the literature, i.e. β-actin and GAPDH. Next, it was determined that a false picture of the expression level of the studied genes can be obtained when a reference gene with variable expression level is used for normalization. RPS17 and 18S rRNA proved to be the most stable reference genes for the WJ-MSC that had been subjected to the lentiviral transfection procedure followed by differentiation. The expression of β-actin and GAPDH was highly unstable and therefore these genes are not suitable for use as reference genes in studies involving WJ- MSC.

Citing Articles

Effects of the Co-Overexpression of the and Genes on the Gamma-Aminobutyric Acid-Ergic Differentiation of Wharton's-Jelly-Derived Mesenchymal Stem Cells.

Borkowska P, Morys J, Zielinska A, Kowalski J Biomedicines. 2023; 11(6).

PMID: 37371846 PMC: 10296716. DOI: 10.3390/biomedicines11061751.

A Strategy for the Selection of RT-qPCR Reference Genes Based on Publicly Available Transcriptomic Datasets.

Nevone A, Lattarulo F, Russo M, Panno G, Milani P, Basset M Biomedicines. 2023; 11(4).

PMID: 37189697 PMC: 10135859. DOI: 10.3390/biomedicines11041079.

Survival and Neurogenesis-Promoting Effects of the Co-Overexpression of and Genes on Wharton's Jelly-Derived Mesenchymal Stem Cells.

Borkowska P, Morys J, Zielinska A, Sadlocha M, Kowalski J Life (Basel). 2022; 12(9).

PMID: 36143442 PMC: 9501059. DOI: 10.3390/life12091406.

Validation of superior reference genes in mouse submandibular glands under developmental and functional regeneration states.

Liu H, He L, Cheng Q, Luo W, Zhao T, Yang D Int J Mol Med. 2022; 50(5).

PMID: 36069228 PMC: 9559174. DOI: 10.3892/ijmm.2022.5188.

Study of the influence of NGF-β gene overexpression in human mesenchymal stem cells on the expression level of SOX1 and neural pathway genes.

Morys J, Borkowska P, Zielinska A, Kowalski J Mol Biol Rep. 2022; 49(6):4435-4441.

PMID: 35348963 DOI: 10.1007/s11033-022-07283-7.

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