siRNA / miRNA gene silencing Human HUVEC IL-8

Get tips on using Stealth siRNA(h)_GATA1 to perform siRNA / miRNA gene silencing Human - LAD2 GATA1

Get tips on using Silencer® Select_HDAC8 siRNA to perform siRNA / miRNA gene silencing Human - UCC HDAC8

Get tips on using Silencer® Select_SPRY2 siRNA to perform siRNA / miRNA gene silencing Human - RMS SPRY2

Get tips on using MEK-1 siRNA (h) to perform siRNA / miRNA gene silencing Human - NHLF MEK1

Get tips on using ERK 1 siRNA (h) to perform siRNA / miRNA gene silencing Human - NHLF ERK1

Get tips on using ENX-1 siRNA (h) to perform siRNA / miRNA gene silencing Human - HaCaT EZH2

Get tips on using Flotillin-1 siRNA (h) to perform siRNA / miRNA gene silencing Human - A2780 FLOT1

Get tips on using Dlx-2 siRNA (h) to perform siRNA / miRNA gene silencing Human - A549 DLX2

Get tips on using SnoA/N siRNA (h) to perform siRNA / miRNA gene silencing Human - SW1990 SnoN

Short hairpin or small hairpin RNA (shRNA) is artificial RNA, which has a hairpin loop structure, and uses inherent microRNA (miRNA) machinery to silence target gene expression. This is called RNA interference (RNAi). These can be delivered via plasmids or viral/bacterial vectors. Challenges in shRNA-mediated gene silencing include 1. Off-target silencing, 2. Packaging shRNA encoding lentivirus, and 3. Stable transduction in cells. RNAi has been designed to have anywhere from 19-27 bs, but the most effective design has 19 bp. In case commercial shRNAs are not available, potential target sites can be chosen within exon, 5’- or 3’ UTR, depending on which splice variants of the gene are desired. One should use the latest algorithms and choose at least two different sequences, targeting different regions, in order to have confidence in overcoming off-target effects. A BLAST search after selecting potential design will eliminate potential off-target sequences. For the second challenge, sequencing the vector using primers for either strand (50-100 bp upstream) is suggested, along with using enzymatic digestion on agarose gel for the vector. Next, once the shRNA-containing vector is packaged in a virus, it is important to check the viral titer before transduction. Finally, using a marker in the lentiviral vector (fluorescent protein or antibiotic resistance), along with qPCR for target gene expression can help in determining the efficacy of transduction and shRNA on its target site.