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Image processing mainly included the following steps: (1) Convert Talairach template [22] into MNI space, and the thalamic subnuclei templates were created using rest software [23].
While there are diverging opinions whether POST or PUT should be used for creating resources in a REST application, our view (supported by [3]) is that this issue is irrelevant for the characterisation of a system as RESTful, as the design goals of the REST software architecture style (scalability, statelessness, cacheability, uniformity) are not violated by either choice.
Relative mRNA expression was calculated using the REST software tool [20].
Relative quantification of the target genes was performed with the well known REST software tool [26].
Relative expression levels were calculated using the Pfaffl method [52], with efficiency correction for each primer set, using REST software [52].
Nonparametric statistical significance of relative RNA expression was calculated with REST software [53] by a pairwise fixed reallocation randomization test with 50,000 repeats.
Hepcidin-1 expression relative to β-actin was compared across groups using both the delta Ct method, and the previously described method [34] using REST software (Qiagen).
Linear detrending and band-pass filtering between 0.01 and 0.08 Hz were performed using the REST software (http://resting-fmri.sourceforge.net/) to partially remove low-frequency drifts and physiological high-frequency noise.
Association with Lim3A levels in embryos was highly significant for this haplotype, by all methods of analysis (Table 3), including pairwise comparisons using REST software (P = 0.0001 for each comparison).
All these analyses were carried out using REST software.
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Samples were analyzed in duplicates and averages were calculated for analysis of the expression ratios using REST-software [ 44].
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