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EF1α is a suitable housekeeping gene for RT-qPCR analysis during osteogenic differentiation of mouse bone marrow-derived mesenchymal stem cells

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Chen X., Zhang B., Zhao Y., Liu P., Zhou Y.

Acta Biochimica Polonica

2013

vol. 60

issue 3

381-386

The expression of predominant housekeeping genes used in RT-qPCR can vary during development and differentiation. The frequently used housekeeping genes (ACTB, GAPDH, 18S rRNA, EF1α and RPL 13a) were evaluated during an early stage of the osteogenic differentiation of mouse bone marrow-derived mesenchymal stem cells (mMSCs) (under normal conditions or treated with CCG-4986) to identify housekeeping genes whose expression remained constant during osteogenic differentiation. When we used RGS4 mRNA, which was determined as copy number per μg of total RNA, to normalize gene expression, we observed that the relative EF1α expression profile was consistent with RGS4 expression after treatment with CCG-4986. All the relative expression profiles of the EF1α, 18S rRNA, and RPL13a housekeeping genes were consistent with RGS4 profiles determined by measuring mRNA copies under normal osteogenic differentiation conditions. The expression profiles calibrated by ACTB and GAPDH were not consistent with those determined using mRNA copy number in untreated cells or cells treated with CCG-4986 under osteogenic differentiation conditions. Under normal osteogenic differentiation conditions, EF1α, 18S rRNA, and RPL 13a are suitable housekeeping genes for RT-qPCR analysis. However, EF1α is the only suitable gene upon CCG-4986 treatment.

Identification and tissue distribution of a novel rat glucocorticoid receptor splice variant

73%

Ju H., Wang X., Liu Z., Liu P., Zhao Y., Xiong R., Zhou Y., Chen X.

Acta Biochimica Polonica

2009

vol. 56

issue 1

109-113

Glucocorticoid receptor (GR) is a steroid hormone receptor that has been shown to play important roles in diverse cellular and physiological processes. More and more evidence has revealed that the effects of glucocorticoids are mediated by the glucocorticoid receptor through genomic or nongenomic mechanisms. A growing number of glucocorticoid receptor splice variants have been identified in human tissues, but few are known in rat tissues. In this work, a novel rGR cDNA, called rGRβ, was cloned from Sprague Dawlay (SD) rat liver. Sequence analysis revealed that the rGRβ mRNA was 39 base pairs (bp) shorter than the rGR mRNA reported earlier. The deleted segment is located in exon 1 and encodes 13 repeated glutamine residues. Both the rGR and rGRβ mRNAs were quantitated by Northern blot hybridization using non-homologous glucocorticoid cDNA probes. Results showed that the rGR and rGRβ mRNAs were most abundant in the lung, the least abundant in the heart, and there were more rGR and rGRβ mRNAs in the kidney than in the liver. The identification of rGRβ may contribute to the understanding of the genomic or nongenomic effects of glucocorticoids.

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2 Acta Biochimica Polonica

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2 Zhao Y.

2 Zhou Y.

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1 Wang X.

1 Xiong R.

1 Zhang B.

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EF1α is a suitable housekeeping gene for RT-qPCR analysis during osteogenic differentiation of mouse bone marrow-derived mesenchymal stem cells

Identification and tissue distribution of a novel rat glucocorticoid receptor splice variant