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Isolation of RNA from blood

Posted by on April 30, 2014

Isolation of RNA from blood

cell-to-dna

 pdf icon2Isolation of RNA

 

  1. 500 µL of fresh blood put on eppendof 1.5 mL
  2. Added 500 µL tri  reagent, was pipetting
  3. Incubated for 5 minutes in romm temperature
  4. Added 100 µL chloroform
  5. shake vigorously for 15 seconds
  6. incubated  for 10-15 minutes at room temperature
  7. Centrifuged at 12,000 rpm 4 ° C for 15 minutes
  8. Taken / transfer aqueous phase ( in top layer ) to another eppendof 1.5 mL
  9. Added 250 µL isopropanol
  10. Incubated for 5-10 minutes at room temperature
  11. Centrifuged 12000 rpm 4 ° C for 8 minutes , the supernatant was discarded , was obtained the white pellet
  12. Added the Pellets with 500 µL 75 % ethanol
  13. Centrifuged 7500 rpm 4 ° C for 5 minutes, the supernatant was discarded completely , was obtained RNA pellet
  14. Briefly air-dry  the RNA pellets for 3-5 minutes remain in the laminar
  15. Added 20 µL of water free RNA
  16. The results are stored at -80 ° C for RT RNA or PCR process , in part to determine the levels and purity using the NanoDrop spectrometer

Note :
a. All procedures must doing in the laminary Air flow
b . All tools (eppendof , yellow type , blue type , white type of must have been in treatment with DEPC water treatment

One Response to Isolation of RNA from blood

  1. Habibi Ainun

    balik lagi ke lab :)

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