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Three approaches were used to normalise the miRNA expression data: global normalisation and endogenous gene normalisation based on RNU44 and RNA48 (Ichimi et al, 2009; Kano et al, 2010).
The results reported in this paper have been obtained by applying a geometric face normalisation based on the eyes positions.
Finally, the ranking of the reduced compound set was obtained using a normalisation based on molecular weight [3] and a consensus scoring in a restrictive way using Chemscore and ASPscore.
DESeq uses a negative binomial distribution to model genic read counts following normalisation based on size factors and variance.
The normalisation based on polyA RNA is similar to the use of the expression levels of all genes for a normalisation in microarray experiments.
Semi-quantitative PCR was performed on different tissues upon cDNA normalisation based on the expression of the actin gene and visualised on agarose gel stained with ethidium bromide.
Similar(49)
(ii) ComBat [7], a Bayesian technique aimed at removing batch effects, and (iii) XPN [8], a cross-platform normalisation technique based on iterative K-means clustering, were also applied for cross-platform normalisation (outlines of the two methods are included in the following paragraphs).
Bond et al gives more details on this method including showing how it is more efficient than traditional normalisation methods based on ratios [57].
The normalisation was based on a kinetic denaturation-reassociation technique described at http://www.vertis-biotech.com/index.php?ip=132.
In addition, we applied a normalisation protocol based on combinations of reference genes determined by geNorm (Vandesompele et al, 2002), as developed for HCC (Supplementary Figure 1).
Calculating a combination normalisation factor based on the geometric mean of three genes for the kairomone experiment and for the growth experiment, stressor-specific regulation of some of the genes involved in basic metabolism is demonstrated.
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