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. 2016 Feb 16;6:20979. doi: 10.1038/srep20979

Figure 3. Splicing analysis of the intronic miRNA expression constructs in mosquito.

Figure 3

(A) The horizontal arrows indicate the positions of the primers used for the expression splicing analysis of the intronic miRNA expression constructs. Amplification with IntronRTF and IntronRTR revealed a 407-bp major band that corresponded to ligation of the NS1 exon, whereas the 540-bp faint band corresponded to the non-spliced transcript. Amplification of rpS7 was used as a loading control. Visible bands were gel purified and sequenced. (B) Plasmids pNS1-GFP-7, pNS1-GFP-210, pNS1-DsRed-7 and pNS1-DsRed-7 transfected into C6/36 cells are shown in the top panel. Recombinant virus VrepNS1-GFP-7, VrepNS1-GFP-210, VrepNS1-DsRed-7 and VrepNS1-DsRed-7 transduced larvae are shown in the bottom panel. If the artificial intron is not removed, the frame-shift mutation in the fused fluorescence reporter will preclude the fluorescence. We detected fluorescence in all of the vector-transfected cells and in the recombinant virus-transduced larvae, confirming that the artificial intron functioned in the mosquito cells.