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The relative expression level of each gene was determined by normalizing to the expression level of the NtEF1 gene in the same sample using the method of REST-MCS as described ( Extraction and quantification of anthocyanin and flavonoid contents in tobacco leaves The processes of extraction, separation and quantification were modified from the previous research ( The freeze-dried tobacco leaf samples were crushed using a mixer mill (MM 400, Retsch, GmbH, Haan, Germany) with a zirconia bead for 1.5 min at a frequency of 30 Hz

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10) Subpart J Exemptions and Variances Finally, Subpart J contains five sections that set forth procedures and requirements for petitioning for a permanent or temporary exemption or variance from any of the proposed requirements

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