Targeting GI-SINE (Growth inducing B2-SINE) RNA in DRG neurons by Antisense Oligonucleotides (ASO)
We identified 2 specific motifs that are enriched in GI-SINE RNA expressed in sciatic-nerve injured DRG neurons compared to non-regulated B2-SINE RNAs. We designed a mix of 5 ASOs (21nt phosphorothioate DNA with LNA-flanks) that target these motifs and transfected those into cultured dorsal root ganglia (DRG) neurons. Control ASO was based on a 21nt non-targetting sequence from shControl backbone (addgene plasmid #85741). Primary DRG neurons were transfected using Dharmafect-4 with ASO at 50nM final concentration 1 hour after plating. Cultures were processed for RNA extraction 48h after transfection for RNA sequencing to identify changes in B2-SINE expression. Overall design: Each replicate (1-4) is based on indepdendent total DRG culture preparation, from 1 mouse in each repeat that was distributed between experimental conditions. Experimental conditions are: ASO control 50nM, ASO mix (GI-SINE ASOs) and untransfected cultures.