siRNA / miRNA gene silencing Human SKOV-3

Get tips on using siRNA RIP3 to perform siRNA / miRNA gene silencing Human - BxPC-3 RIP3

Get tips on using Accell Human VDAC1 (7416) siRNA - Set of 4 to perform siRNA / miRNA gene silencing Human - PC-3 VDAC1

Get tips on using PACE4 siRNA (h) to perform siRNA / miRNA gene silencing Human - PC-3 PACE4

Get tips on using siRNA TRAF6 (h) to perform siRNA / miRNA gene silencing Human - Calu-3 TRAF6

Get tips on using SignalSilence® FoxO3a siRNA to perform siRNA / miRNA gene silencing Human - Calu-3 FOXO3

Get tips on using SignalSilence® FoxO1 siRNA to perform siRNA / miRNA gene silencing Human - Calu-3 FOXO1

Get tips on using SMARTpool: siGENOME PAK4 siRNA to perform siRNA / miRNA gene silencing Human - OVCAR-3 PAK4

Get tips on using siRNA Junction plakoglobin (SASI_Hs01_00246520) to perform siRNA / miRNA gene silencing Human - BxPC-3 plakoglobin

Get tips on using Oct-3/4 siRNA (m) to perform siRNA / miRNA gene silencing Mouse - P19 Oct4

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.