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Problems with extraction of RNA from primary culture


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#1 Mettiu

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Posted 15 January 2013 - 09:00 AM

Hi there,

I am working with mixed glial/neuronal primary culture (P1&P2) from cortex. I am having some problems with the extraction of RNA.
From 2 X 10^6 cells per well I can't get more than 70ng/ul. I have used kits like Qiagen, Roche, and TRIZOL.

Does anybody have any suggestion?

Thanks

#2 Trof

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Posted 15 January 2013 - 09:35 AM

Are you aware that neuronal tissue has high lipid content? Did you use specific kits or modifications for this kind of tissue?

(By the way, DON'T post multiple identical topics into several subforums)

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#3 Mettiu

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Posted 16 January 2013 - 05:40 AM

Hi,

Thanks for replying. I have used specific kits for cell:
Qiagen mini kit
Roche high pure RNA tissue kit
TRIZOL, following a protocol from other colleagues that are used to work with primary cultures and cell cultures in general

#4 Trof

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Posted 16 January 2013 - 07:30 AM

There is a special Qiagen kit for lipid tissues RNeasy Lipid Tissue kit, but Trizol isolation protocol should be suitable even for those.

Our country has a serious deficiency in lighthouses. I assume the main reason is that we have no sea.

I never trust anything that can't be doubted.

'Normal' is a dryer setting. - Elizabeth Moon


#5 Mettiu

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Posted 21 January 2013 - 02:41 AM

Hi,

I haven't tried that one. The last one that I have used is ReliaPrep RNA Cell 20Miniprep - Promega. This one works really well.

Cheers




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