DNA contamination in microRNA sample - How to remove the genomic DNA from microRNA sample? (May/08/2007 )

Dear all,

I used the miRNeasy mini kit to prepare the microRNA. However, I found DNA contamination occured in my microRNA sample. How can I remove the DNA? Would you kindly to give me a protocol for the removal of DNA from microRNA?

Thank you very much!!!!!

siuchi

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use turbo free DNAse from ambion. It's amazing. I use it for my RNA preparations checked on agilent and it's reliable.

NB : no financial intereest in it

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Yea, agreed with fred_33. Use Dnase to remove any trace of DNA. Then you have to clean that up. Phenol chloroform method?

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Dear fred_33 and timjim,

Thank you so much!!!!!
I still have a question: How to purify the miRNA after the DNase digestion?

Many thx!!!

siuchi

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turbo DNAse is sold as enzyme linked to agarose beads.
so assuming a 60µl reaction volume, i pellet the beads by spinning and pipett 50µl (55 if the sample is REALLY low concentrated or REALLY needed).
Don't pipett more than 50 if you wanna perform RTPCR

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I am not too sure how to purify miRNA after the digestion. You might need some extra or extensive protocol to do it.
I dealed with normal mRNA, that's why I just use the normal phenol chloroform method.

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