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Quantitative, two-colour Western blot analysis was performed on three biological repeats using a LICOR Odyssey FC instrument and Image Studio V2.0 analysis software.
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Somites from 20-25 embryos were pooled for western blot analysis; three biological repeats used material from independent experiments.
Briefly, three biological repeats were executed using a SYBR Green real-time PCR Master Mix TOYOBOO) using ABI 7500 Fast Real-Time PCR System (Applied Biosystems).
miRNA real time PCR was performed on 2 additional biological repeats (total of 5 repeats) using TaqMan specific Small RNA primer and probe sets for miR-181c, miR-411, miR-659, miR-510, miR-126, miR-15a and miR-20b (Applied Biosystems, Inc., Foster City, CA, USA).
Three biological repeats were used for each treatment.
For each drug treatment, 3 biological repeats were used, and each experiment was repeated 2 3 times.
For all qPCR analysis at least three biological repeats were used unless stated otherwise.
Only the phosphopeptides with unambiguous (class I) phosphorylation sites identified from three biological repeats were used for further analysis.
Mean values of three biological repeats were used to plot figures, and error bars on each figure represent the standard errors.
P-values in the experiment without biological repeats were calculated using DEGseq software.
The average colony area percentage from three independent biological repeats was calculated using the 'ColonyArea' ImageJ plugin (Guzmán et al., 2014a).
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