Immunohistochemistry Wilms Tumor 1 (WT1) Rabbit

Get tips on using ApopTag® Peroxidase In Situ Apoptosis Detection Kit to perform TUNEL assay cell type - HNSCC Detroit 562 human head and neck tumor cells

Get tips on using Human NRG1-beta 1/HRG1-beta 1 DuoSet ELISA to perform ELISA Human - NRG1

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.

Get tips on using HyClone Dulbecco's Modified Eagle Medium (DMEM)/ F12 1:1: Liquid to perform 3D Cell Culture Media U87MG cells- glioblastoma spheres

Get tips on using HyClone Dulbecco's Modified Eagle Medium (DMEM)/ F12 1:1: Liquid to perform 3D Cell Culture Media hiPSC-derived cortical organoids

Get tips on using HyClone Dulbecco's Modified Eagle Medium (DMEM)/ F12 1:1: Liquid to perform 3D Cell Culture Media hiPSC-derived forebrain organoids

Get tips on using HyClone Dulbecco's Modified Eagle Medium (DMEM)/ F12 1:1: Liquid to perform Stem cell Differentiation media hDPSCs differentiation into adipogenic cells

Get tips on using HyClone Dulbecco's Modified Eagle Medium (DMEM)/ F12 1:1: Liquid to perform Stem cell Differentiation media hDPSCs differentiation into osteogenic cells

Get tips on using Beclin-1 Antibody to perform Autophagy assay cell type - THP 1

Gene silencing through the use of small interfering RNA (siRNA) has become a primary tool for identifying disease-causing genes. There are several aspects for preparing and delivering effective siRNA to knockdown a target gene. The length of siRNA should be 21–23nt long with G/C content 30–50%. If a validated siRNA sequence for your target gene is not available, use siRNA generated against the entire target gene ORF. Always work with two or three different siRNA constructs to get reliable results. If you are not sure how much siRNA to use for a given experiment, start with a transfection concentration of 10-50 nM and use siRNA-specific transfection reagent to ensure efficient siRNA delivery in a wide range of cells.