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Sample (dissected frozen brain region)

Data generation. Sample (dissected frozen brain region). RNA extraction (total RNA) QIAZOL and RNeasy mini kit. RNA quantification (Bioanalyser). Reverse transcription MMLV RT-ase Random priming. Q-PCR with gene-specific primers Multiple reference genes Genes of interest.

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Sample (dissected frozen brain region)

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  1. Data generation Sample (dissected frozen brain region) RNA extraction (total RNA) QIAZOL and RNeasy mini kit RNA quantification (Bioanalyser) Reverse transcription MMLV RT-ase Random priming Q-PCR with gene-specific primers Multiple reference genes Genes of interest Data analysis using raw Ct values Calculate geometric mean of 3 reference genes Express crossing threshold of each sample with respect to the least expressed sample for the geometric mean of reference genes (DCt) Express crossing threshold of each sample with respect to the least expressed sample for each gene of interest Calculate DCt (sample) – DCt (reference) = DDCt Calculate expression ratio according to the formula 2-DDCt

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