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In order to search for those transcripts showing similar expression fluctuation in both pericarp tissues, a similar analysis (5% FDR in a 6-class Limma and 2-fold change) was run considering skin and flesh samples in the same time point as replicates.
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Fruit flesh samples were immediately frozen in liquid nitrogen and kept in −80 °C until use.
Older laboratory techniques to identify fish meat looked at the mix of proteins in flesh samples, but were unreliable, expensive and cumbersome.
Starch content in flesh samples of freshly harvested tubers was here estimated at ~14.5%% of total fresh weight for the three tested lines (anova; P = 0.594, F = 0.569), comparable to starch levels observed earlier in tubers of the same cultivar [ 26, 38].
More Gly accumulated in the outer-canopy peels of 'Gala' and 'Mutsu' fruit, whereas no differences were observed in the flesh samples.
Although the peel of outer-canopy fruit had more myo-inositol than that of inner-canopy fruit, no obvious positional effects were found in the flesh samples.
The analysis of free amino acids was performed as described by Zhang et al. Frozen apple peel and flesh samples (0.5 g) were ground in 1.5 mL of 70% methanol containing 2% formic acid at 0 4 °C and then centrifuged at 10 000 g for 10 min at 4 °C.
The same cut-offs were applied in Limma after considering Tempranillo skin and flesh samples for the same time point as replicates to identify transcripts with consistent oscillation in expression along the pericarp.
In this case, Tempranillo skin and flesh samples for the same time point were considered replicates to pick out transcripts with consistent expression along Tempranillo pericarp that was the tissue analysed in Verdejo.
The hybridization signal for VvMybA1 and VvMybA3 microarray probe sets was constitutively higher in Tempranillo berry skin and in Verdejo pericarp when compared to Tempranillo flesh samples (Additional file 1).
Flesh samples were taken by cutting two wedges, each approximately one-eighth of the fruit.
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