Dear All,
mirna primer showing some problem in the melting curve? any idea why? As attached is the melting curve. The forward sequence is obtained from miRBase and reverse primer is universal.
It looks like a problem with primer-dimers and an unspecific product. You can run the reaction on an agarose gel to see what you have there. Have you check with primer blast whether your primers are specific?
Samantha Filipów Hi, thanks for your response. Yeah, I already run gel electrophoresis, three bands in total.. how can I troubleshoot? can we blast miRNA primer?
https://www.ncbi.nlm.nih.gov/tools/primer-blast/ put sequences of your primers, choose RNAseq database and the correct species, to see if your primers are specific and if they are complement to each other, giving primer-dimers. Did you run any controls for your qPCR? gDNA?
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