Hi,
During RNA isolation, why is it required to spin the tubes not more than 10000 rpm rather than 12000 rpm after adding "isopropanol"?
Thanks!
Hi,
During RNA isolation, it's important to spin the tubes at a specific speed after adding isopropanol to ensure that the RNA remains intact and undamaged.
The reason for spinning the tubes at 10,000 rpm rather than 12,000 rpm is because at high speeds, the centrifugal force can cause mechanical shearing of the RNA, which can lead to degradation and loss of the integrity of the RNA. This is especially true when the sample is subjected to high forces, like the ones present during a 12000 rpm spin.
Additionally, spinning at lower speeds can minimize the formation of foamy layers which can cause the RNA to get entrapped and degrade as well.
In general, it is recommended to spin samples at the lowest speed that can still effectively pellet the desired components, as this will help to minimize the potential damage to the RNA or other fragile molecules present in the sample.
It is worth noting that in some protocols, the spin speed might be adjusted based on the volume of the sample, the type of centrifuge used or even the type of RNA isolation kit used. It's always best to check the instructions provided with your specific kit or consult the literature for recommended spin speeds for your samples and equipment.
Regards.
First of all, I doubt there is really much difference between spinning at 10K or 12K RPM. In fact, listing the speed alone is meaningless since the centrifugal force being applied is a function of speed and distance from the axis (rotor radius). So a larger rotor (think a 24 place microfuge) at 10K will actually generate much more centrifugal force than a smaller rotor at 12K.
Michael J. Benedik
I agree with you that there is not much difference in 10K or 12K, however, in this case if there was difference, it would be relative. Since, I guess the person will use the same centrifuge at 12K which will be stronger as compared to 10K.
Also, I do not agree much with the explanation above from Mohammad Alzeyadi as centrifuge is not a vortex which can sheer the molecule. I guess the whole answer is a misinterpretation of the question. Isopropanol help DNA/RNA precipitate faster, so it is recommended to use everything less or milder (time for incubation/precipitation, temperature[higher] and time) as compared to ethanol to avoid precipitation of salts.
Additionally, as Michael J. Benedik said, just mentioning centrifuge speed is meaningless without knowing the other parameters.
Avoid damage to RNA, because high speed may cause mechanical damage to RNA.
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