Hello,
I have a strange problem.
In my lab we are trying to use CRISPR/Cas9 technique to create a Knock-out of a miRNA in Jurkat cells ( T cells) .
In detail I used a crRNA that target specifically the miRNA and I have used a DONOR DNA of 64bp.
From sequencing I see the presence of the DONOR in both the alleles, so the sequence was effeciently mutated.
To confirm this data I have done a qPCR of this miRNA, and unfortunally I see yet the presence of this miRNA, how is this possible?
I don't understand, because the Donor break the miRNA sequence, and the result of sequencing is clear.
In my Jurkat cells there isn't the wild type allele, but only mutated ones.
Could you help me, please?
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