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This study was designed to (i) identify stable reference genes for the analysis of gene expression during in vitro differentiation of rat adipose stromal cells (rASCs), (ii) recommend stable genes for individual treatment conditions, and (iii) validate these genes by comparison with normalization results from stable and unstable reference genes.
The application of geNorm with default limit (M < 1.5) ruled out unstable reference genes and left minimal number of suitable reference genes for each group in the end.
Extensive variation in C t values is to be expected in a non-normalized data set, and so the algorithm may not be able to effectively distinguish between stable and unstable reference genes.
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The reviewers were referring specifically to the inhibitor fostriecin, which is somewhat notorious in being unstable (see the reference in the manuscript to the Swingle et al. review).
However it is found that the reference trajectory is unstable throughout the deployment and a linear feedback control is introduced for stabilization.
Prior to initiation of this study, we performed preliminary experiments using GAPDH and β-Actin and found that these reference genes were unstable in these inflamed tissue samples.
Several effects can distort the abundance of RNA transcripts during extraction and preparation before RNA analytics using, e.g., microarrays: The first problem concerns the degradation of the RNA in vitro [ 1- 4]: The quality of purified RNA is variable and after the extraction during storage rather unstable (see [ 5] and references cited therein).
This reference model is locally unstable in most operating points, making open loop simplification impractical.
The unstable points are obtained from the reference by reducing significantly the equivalence ratio and by an important dilution by nitrogen respectively.
There are references to his being psychologically unstable and to a recent suicide attempt.
For balanced and unbalanced loads the VIQ PPS is stable where a reference iSLIP single-plane switch is unstable.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com