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. 2017 Nov 3;8(11):303. doi: 10.3390/genes8110303

Figure 5.

Figure 5

In silico analysis of the 200 bp fragment sequence from the NADHox gene for the formation of small RNAs and their detection by stem loop RT-PCR. (A) In silico prediction of siRNAs from the 200 bp fragment, using the online tool (bioinfo.clontech.com/rnaidesigner/sirnaSequenceDesignInit.do. (B) Predicted secondary structure of the ssRNA (200 bases) obtained with the MFold Web Server, indicating the position of a putative miRNA. (C) Structure of the Stem-loop NADHox primer obtained with the Universal Probe Library probe #21 hybridizing to the target siRNA. (D) RT-PCR amplified fragments of the stem-loop probe with the captured siRNA run on a 16% polyacrylamide gel: Lane 1, siRNA amplification with a size of ~53 bp, using primers siRNA NADHox Fw and UniLoop; Lane 2, amplification of the stem-loop region (stem-loop NADHox primer) with the specific siRNA #4 (or miRNA) showing the expected size of 63 bp; M, O’RangeRuler 10 bp DNA Ladder (Thermo Scientific).