rna-isolation-purification-tissue-mouse-cornea

Get tips on using Brilliant Violet 510™ anti-mouse CD69 Antibody to perform Flow cytometry Anti-bodies Mouse - CD69

Get tips on using PerCP-Cy™5.5 Hamster Anti-Mouse CD69 to perform Flow cytometry Anti-bodies Mouse - CD69

Get tips on using PE-Cy™7 Hamster Anti-Mouse CD11c to perform Flow cytometry Anti-bodies Mouse - CD11c

Get tips on using Mouse PAI-1 total antigen assay ELISA kit to perform ELISA Mouse - Serpin E1/PAI-1

Get tips on using Mouse/Rat IGF-I/IGF-1 Quantikine ELISA Kit to perform ELISA Mouse - IGF-I

Get tips on using EndoFree Plasmid Mega Kit (5) to perform DNA isolation / purification Plasmid purification

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 Mouse/Rat Osteopontin (OPN) Quantikine ELISA Kit to perform ELISA Rat - OPN

Get tips on using Mouse/Rat Angiopoietin-2 Quantikine ELISA Kit to perform ELISA Rat - ANGPT2

Get tips on using Purified anti-mouse/rat/human FOXP3 Antibody to perform Western blotting FOXP3