siRNA / miRNA gene silencing Human Primary Human Hepatocytes

Get tips on using Slc1a2 to perform siRNA / miRNA gene silencing Rat - Glial cells GLT-1

Get tips on using mirVana™ miRNA Isolation Kit, with phenol to perform RNA isolation / purification Cells - primary human pulmonary artery smooth muscle cells

Get tips on using Glut1 siRNA and shRNA Plasmids (h) to perform siRNA / RNAi /miRNA transfection Human Cells - HT-1376 GLUT1

Get tips on using CD74 siRNA and shRNA Plasmids (h) to perform siRNA / RNAi /miRNA transfection Human Cells - HT-1376 CD74

Get tips on using MicroRNA isolation kit to perform siRNA / miRNA gene silencing Rat - IEC-6 HuR

Get tips on using Rock-2 siRNA and shRNA Plasmids (h) to perform siRNA / RNAi /miRNA transfection Human Cells - HT-1376 ROCK2

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 have 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 efficacy of transduction and shRNA on its target site.

Get tips on using FlexiTube GeneSolution GS27279 for Tnfrsf12a to perform siRNA / miRNA gene silencing Mouse - B16-BL6 FN14/Tnfrsf12a

Get tips on using MammoCult™ Human Medium Kit to perform 3D Cell Culture Media Primary human breast tumors-Mammospheres

Get tips on using 3D-Gene® Mouse miRNA Oligo chip (ver.21) to perform Microarray Gene expression arrays - Mouse liver tissue Cyanine-3-CTP