Dear All,
In AAV vector if we clone a miRNA we clone this pri-miRNA sequence in to the 5' of the reporter gene.
For example this is the empty vector : https://www.addgene.org/21893/
and this is the miRNA cloned version: https://www.addgene.org/21894/
the pre-miRNA is cloned within the EFa intron which is located in the 5' of reporter gene.
Onces the mRNA is transcribed it will contain miRNA loop in the 5' of the reporter gene and it will be cleaved by the drosha to process miRNA. Meanwhile reportergene will lose its 5' cap due to cleave. But in articles this model is widely used and it works.
so my question is that how does this model work? 5' cap should be essential for ribosomal localization.
The picture from the paper "Therapeutic microRNA Delivery Suppresses Tumorigenesis in a Murine Liver Cancer Model" explains the cloning orientation.
Thanks
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