RNA extraction - (Dec/01/2006 )
Do we have to use "freshly prepared 70% ALCOHOL" while doing RNA extraction? I did an expet. with about 1.5 million cells X 4 samples and all four gave very low RNA content. Thinking back-I used 70% alcohol prepared 3 weeks back. Could that be the problem?
Since you are using the RNeasy kit, I think it is the amount of cells that you put it. Check the instruction, it mentioned about amount of cells needed. If too much, then your RNA level will be low. If too little, same thing will occur. Good luck.
better check for RNase contamination too.
now i am using a RNA extraction kit from GE health care. its very good, easy and got high yield. takes about 1 hour to extract RNA from 6 samples
80% in ready to use aliquots in microcentrifuge tubes, store at -20C... works for me.
For EtOH precipitation on DNA you use ice-cold EtOH: my advice is to do the same with RNA. Works like a charm.
i don't prepare new 70% ethanol. but i don't use the same bottle for other purpose (RNA work only). i seal the cap using parafilm and store in fridge.
it works fine for me.
i make 80% ethanol once and use it upto finish it (may be upto 3 months)....no problem yet
I also think that you do not need to prepare fresh ethanol....
RNA is a "weaker" thatn DNA, so you need to work in a very clean space , using solutions meant only for that porpoise.
If you do this, there shouldn't be any problem....