How is detecting miRNAs different from detecting normal mRNA transcripts with the qNPA ArrayPlate?
miRNAs are significantly shorter (typically 21 to 23 bases) than mRNA and cannot protect the standard 50 nucleotide-long DNA oligo used for mRNA detection during the S1 nuclease detection step. Therefore, we have designed smaller qNPA protection oligos to detect these miRNA sequences. These miRNA-targeted qNPA protection oligos cannot support hybridization of the Detection Linker oligonucleotide due to their short length. Therefore, these qNPA oligonucletides are biotinylated to facilitate subsequent detection. An avidin-HRP conjugate is used to detect miRNA hybridization instead of the Universal Detection Linker used in the standard protocol.
