Doubt: Trizol protocol
Posted 01 December 2009 - 06:52 AM
1) It has been told to not to take tissue more then 10% of the volume of Trizol reagent used. Why? What would happen if more tissue is used ?
2) After isopropanol precipitation, to harvest the RNA pellet it is stated to not to exceed 12,000 g speed. Why?
3) "Do not dry the RNA by centrifugation under vaccum" Why?
Posted 01 December 2009 - 07:19 AM
2. Too compact a pellet for resuspension? Break the tubes? Precipitate other stuff like proteins?
3. Totally dry DNA or RNA is very difficult to resuspend, taking many days.
Posted 02 December 2009 - 02:35 AM
Posted 02 December 2009 - 05:22 AM
2. I do the precipitation and washes at max speed (14k g)...never had a problem, so I wonder why they say it!! maybe as phage said, to avaoid compaction of the pellet!
3. I'd read it as "Dont vacuum dry the RNA for too long" (I do a 3 minutes vacuum spin after the ethanol wash and resuspend the pellet asap after making sure there is no more ethanol leftover)
Posted 03 December 2009 - 02:35 AM
Posted 03 December 2009 - 05:54 AM
Posted 03 December 2009 - 07:08 AM
Posted 04 December 2009 - 01:38 AM
Posted 04 December 2009 - 04:55 AM
Posted 05 December 2009 - 04:36 AM