siRNA / miRNA gene silencing Human SUIT-2

Get tips on using Timp2 siRNA to perform siRNA / miRNA gene silencing Mouse - BV2 TIMP-2

Get tips on using siGENOME Human MINK1 siRNA to perform siRNA / miRNA gene silencing Human - RMS MINK

Get tips on using siGENOME Human MAP4K2 siRNA to perform siRNA / miRNA gene silencing Human - RMS MAP4K2

Get tips on using siGENOME Human PAK1 siRNA to perform siRNA / miRNA gene silencing Human - HeLa PAK1

Get tips on using ON-TARGETplus human ATG16L1 siRNA to perform siRNA / miRNA gene silencing Human - SHSY5Y ATG16L1

Get tips on using ON-TARGETplus Human TET1 siRNA to perform siRNA / miRNA gene silencing Human - A172 TET1

Get tips on using ON-TARGETplus Human ARL2BP siRNA to perform siRNA / miRNA gene silencing Human - HeLa BART/ARL2BP

Get tips on using ON-TARGETplus Human BECN1 siRNA to perform siRNA / miRNA gene silencing Human - U251 Beclin 1

Get tips on using Silencer® GAPDH siRNA (human) to perform siRNA / miRNA gene silencing Human - Jurkat GAPDH Lipid

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.