Erickson:REal-time PCR

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Isolation of Total RNA from Fleas

Master Mix:
• 5μL 2x SYBR green
• 1μL Forward primer (5μM)
• 1μL Reverse primer(5μM)
• 3μL PCR grade water

  • Note: This is for a single reaction, multiply by number of reactions needed


Procedure

     1. Add 1μL of DNA or cDNA teplate to 9μL Master Mix in a 96-well Multiwell plate.
    2. Seal the multiwell plate with Light Cycler 480 Multiwell sealing foil
    3. Centrifuge the Multiwell plate at 15xg for 2 minutes
    4. Transfer the Multiwell plate to the holder of the Light Cycler and close(push button to open and close)
    5. Start the PCR program