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Division I.4.4 Sample preparation for RT-PCR and RT-qPCR based surveillance programmes

United Kingdom

Original text. Later amendments are not shown.

If samples were collected in transport medium, the procedure set out in points I.4.2 and I.4.3 shall be carried out. After centrifugation, supernatant shall be collected and RNA extracted. If further examination is not to be undertaken directly after centrifugation, the samples shall be immediately frozen at – 20 °C or below.

For the analysis of fish tissues preserved in RNA stabilization reagent, subsequent work shall be carried out within the following time scales for samples stored at different temperatures:

  • samples stored at 37 °C: 1 day;

  • samples stored at 25 °C: 1 week;

  • samples stored at 4 °C: 1 month;

  • samples stored at – 20 °C: indefinitely

Pooled samples in RNA stabilization reagent shall be treated like single samples in RNA stabilization reagent. For samples pooled in RNA stabilization reagent, the sample amount shall not exceed that recommended by the manufacturer for extraction with RNA kits, such as RNeasy Mini kits (Qiagen) or similar. If larger samples are pooled, the extraction kits or methods must reflect this pooling.

Samples collected in RNA stabilization reagents shall not be used for cell cultivation.