Any suggestions for high quality miRNA extraction would be appreciated.
Hi Hanie,
I've used the specially designed Qiagen miRNA kits before with good results to perform qPCR measurement of miRNAs further down the line, machery nagel also produce some cheaper kits however with lower yield obtained in my experience.
Dear Steven Dodd
Thank you for such detailed responses.
I really appreciated you taking the time to answer my question.
Dear Hanie Motahar
Pls refer to the following article which explained deeply how to isolate miRNA:
doi.org/10.1073/pnas.1117207108
The used kit is: miRNeasy Mini kit (Qiagen).
They stated the following:
Serum miRNA isolation and quantitation. Serum was treated with 5 volumes of QIAzol (Qiagen) for the denaturation of protein contents and subsequent isolation of RNA. Due to the absence of a known housekeeping microRNA (miRNA) in human serum, 25 fmol of synthetic cel-miR-39 (5′-UCACCGGGUGUAAAUCAGCUUG-3′) was added as a means of normalization (9). After adding 1 volume of chloroform, aqueous and organic phases were separated by centrifugation at 12;000 × g × 15 min at 4 °C. The aqueous phase was treated with 1.5 volumes of 100% ethanol and loaded to a miRNeasy Mini column for RNA extraction. After extensive washes, purified RNA was eluted from the column membrane with 50 μL of nuclease-free water and stored at −80 °C until analyzed.
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