Validation of Reliable Reference Genes for Real-Time PCR in Human Umbilical Vein Endothelial Cells on Substrates with Different Stiffness
作者： Gan Chen , Lian Zhao , Jiantao Feng , Guoxing You , Quanmei Sun
DOI: 10.1371/JOURNAL.PONE.0067360
关键词:
摘要: Background The mechanical properties of cellular microenvironments play important roles in regulating functions. Studies the molecular response endothelial cells to alterations substrate stiffness could shed new light on development cardiovascular disease. Quantitative real-time PCR is a current technique that widely used gene expression assessment, and its accuracy highly dependent upon selection appropriate reference genes for normalization. This study aimed evaluate identify optimal use studies stiffness. Methodology/Principal Findings Four algorithms, GeNormPLUS, NormFinder, BestKeeper, Comparative ΔCt method, were employed nine candidate genes. We observed stability potential varied significantly human umbilical vein substrates with different stiffness. B2M, HPRT-1, YWHAZ are suitable normalization this experimental setting. Meanwhile, we normalized YAP CTGF using various demonstrated relative quantification according genes. Conclusion/Significance: Consequently, our data show first time set stably expressed accurate exploring effect subendothelial matrix stiffening cell function. furthermore caution against GAPDH ACTB setting because low study.
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