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After the needle punctures the stopper, the vacuum may naturally draw some or all of the bacteriostatic water into the vial with little additional pressure

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Real time quantitative PCR Total RNA was obtained from 1.5 mL of aliquots from endpoints Z, A and B, using the QIAamp RNA Blood kit (Qiagen, Hilden, Germany), and cDNA synthesis was performed from 2 g of total RNA using the High-Capacity cDNA Reverse Transcription Kit (ThermoFisher, Waltham, Massachusetts), both according to manufacturers protocols

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