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RNA extraction without cell disruption

RNA extraction without cell disruption - Molecular Biology Techniques

RNA extraction without cell disruption - Molecular Biology Forum. Includes forums for common molecular biology techniques.


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Old 02-06-2013, 01:01 PM
Pipette Filler
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Default RNA extraction without cell disruption



Hi,
I need to extract total RNA from filters containing bacterial cells from marine samples. I am just starting with this and am afraid to waste my filters with inefficient extraction methods. There is an established protocol for DNA and RNA extraction from such filters in our lab, and it involves cell dirsruption with glass beads and phenol-extraction. However the typical RNA-yield is usually rather low.

Just recently a collegue forgot the beads, and simply vortexed the filter fragments in a phenol solution. suddenly the yield was about 3-4 times higher! Does this even make sense?

Could it be that phenol-extraction and rigorous shaking is enough to extract RNA from most cells, and that mechanical cell disruption is contraproductive (eg. because the extracted RNA gets sheared and becomes too small to be efficiently recovered by alcohol precipitation?).

Or is it just that our established protocol is to rigourous (too many beads and to strong shaking), and we should be less brutal to the samples?

Has anybody got similar experiences? What do you think?
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