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RNA extraction from trout embryos


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#1 tams

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Posted 08 July 2010 - 03:17 AM

Hi,
I've been having problems trying to extract RNA using TRI reagent from early stage trout embryos which have very small amounts of RNA and lots of lipid/protein. The problem seems to be that I get such small amounts of RNA that protein and phenol contamination becomes very evident causing very low 260/280 and 260/230 ratios. I've tried a high salt solution modifcation (NaCl and sodium citrate) of the isopropanol precipitation step and an additional ethanol/sodium acetate wash at the end of the protocol and both of these seem to remove some of the protein/phenol contamination, but also reduce my RNA yield further so the overall quality is still very bad.
If anyone has any ideas of anything else I can try that would be great!
Thanks

#2 mdfenko

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Posted 08 July 2010 - 07:12 AM

you can remove phenol with chloroform (or chloroform/isoamyl alcohol).

Edited by mdfenko, 08 July 2010 - 07:12 AM.

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#3 apbrown

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Posted 08 July 2010 - 11:02 AM

I have a question regurading RNA extraction using the nano drop how do I determine weither or not my RNA is good where should the peak be between.

#4 tams

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Posted 09 July 2010 - 03:19 AM

you can remove phenol with chloroform (or chloroform/isoamyl alcohol).


Thanks- I will give it a go.




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