Yes..DEPC-water or I usually buy ultrapure water from Invitrogen...and the pellet goes back into solution fine as long as it's not over dried. As far as OD is concerned, it's always worked fine...with ratios from 1.6 to 1.8, and for OD readings, (I will dilute into TE and blank with TE as pure water gives inconsistent readings). Usually not concerned since I'm doing a Northern or RT, so it gets internally controlled.
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#17
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Posted 19 August 2010 - 06:05 PM
An update:
After changing everything, I tried to isolate some fresh samples. It turns out that it was my sample buffer--there must have been something in it that was degrading the RNA. After making fresh sample buffer, I saw three very pretty RNA bands in my sample on the agarose gel, plus OD readings of 1.7. Additionally, I took some older RNA and was able to see bands.
Thanks all for your advice. Everyone provided very useful comments!
-b
After changing everything, I tried to isolate some fresh samples. It turns out that it was my sample buffer--there must have been something in it that was degrading the RNA. After making fresh sample buffer, I saw three very pretty RNA bands in my sample on the agarose gel, plus OD readings of 1.7. Additionally, I took some older RNA and was able to see bands.
Thanks all for your advice. Everyone provided very useful comments!
-b
Edited by btc8, 19 August 2010 - 06:06 PM.
#18
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Posted 20 August 2010 - 07:08 AM
Ah. I thought that you were extracting directly from the dish using trizol...
#19
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Posted 21 August 2010 - 01:48 PM
NemomeN007, on 20 August 2010 - 07:08 AM, said:
Ah. I thought that you were extracting directly from the dish using trizol...
I usually do this. I used to trypsinize and spin down to pellet, but I found that this would decrease my yield.
