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A random selection of samples covering all conditions and tissues was used to determine the most stable endogenous control genes.
We demonstrated earlier that TLDA included the most stable endogenous controls, and, considering the suggestion of others [21], [22], we used the results from TLDA as a standard to evaluate the performance led by different normalization methods when applied to the analysis of LNA and beads arrays.
While 18S has been reported as the most stable endogenous control under various experimental conditions [15], [36], including omental and subcutaneous adipose tissues from obese and diabetic patients [15], others have noted modest variability in 18S following hormonal challenge of cultured human adipocytes [2].
GAPDH was selected as the most stable endogenous control gene.
RSP16 was selected as the most stable endogenous control gene.
SASqPCR [ 17] was used to identify the most stable endogenous control genes.
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While our studies highlighted 18S as the most stable of endogenous controls under all four of our defined experimental conditions, it should be noted that there are concerns with using rRNA as a control which may not be influenced by degradation machinery in a matter similar to mRNA [15], [37].
For analysis of miRNA by qRT-PCR, RNU44 and RNU6B controls were examined to determine the most stable small RNA endogenous control.
These genes are reported within the most stable set of endogenous control genes.
ACTB and GAPDH were selected as the most stable control genes from four endogenous genes tested (GAPDH, ACTB, HPRT1 and BCR) using the geNorm algorithm.
GAPDH was selected as endogenous control as it proved most stable, which is consistent with the literature [50].
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