Isolated RNA

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biology_06er
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Isolated RNA

Post by biology_06er » Wed Mar 26, 2008 12:32 am

Hi there,

We did an experiement last week where we isolated RNA from rat liver...the gel shows 2 distinct bands--just wondering if these two bands are present representing the diff. subunits like 28S etc...this means that the experiement was successful right?
And if the gel shows "smearedness" this means the experiment was ruined right?, due to contamination of Rnases causing the RNA to degrade...

Thanks,
b_06er

Cat
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Post by Cat » Wed Mar 26, 2008 12:39 am

It depends on what you did. If you isolated total RNA and ran it directly, you suppose to get a smear. If you did RT-PCR and ran that on the gel, you would expect a product of a particular size…

biology_06er
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Re: Isolated RNA

Post by biology_06er » Wed Mar 26, 2008 1:07 am

it looks like the "intact" lane on the following website

http://www.ambion.com/techlib/append/supp/rna_gel.html


as for how we ran it..the lab technicians did it for us...we only loaded it into the wells and left it with them!... :shock: ...and looking thru my instructions it doesn't mentioned RT-PCR or if done directly but i guess from the result it was RT-PCR??

Thanks,
b_06er

Cat
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Post by Cat » Wed Mar 26, 2008 3:45 am

Arhh… Now I understand what you are talking about. I was under impression you were talking about two bands without smear vs. smear.

A more common gel result (but then again it depends on amount of RNA loaded):

http://www.zyagen.com/Products%20files/ ... _small.gif

If your smear has distinct band for rRNA subunits then it is intact (if it doesn’t – it’s degraded). So you were right.

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