InvivoGen provides a plasmid-based system developed to knockdown efficiently the expression of a wide variety of mammalian genes. This system represents a simple and affordable method to generate siRNAs by eliminating the need to synthesize RNA oligonucleotides. The key element of this system is psiRNA, a RNA polymerase III based expression vector that enables the endogenous production of small double-stranded RNAs for gene silencing applications. Furthermore, psiRNA can be stably transfected into a given cell line to achieve persistent down-regulation of a target gene.

Description The psiRNA™ system consists of the following components: psiRNA™ G2 cloning vectors E.coli GT116 strain Sequencing primers E. coli Fast-Media® psiRNA G2 Vectors A new generation of psiRNA plasmids is now available. See description on the psiRNA Vectors page. E. coli GT116 Strain Hairpin structures are known to be unstable in E. coli due to their elimination by a protein complex called SbcCD that recognizes and cleaves hairpins [1]. To increase their stability in E. coli, we developed GT116, a sbcCD mutant strain that significantly improves the number of recombinant clones. GT116 genotype: F- mcrA D(mrr-hsdRMS-mcrBC) f80lacZM15 DlacX74 recA1 endA1 DsbcC-sbcD. LyoComp GT116  GT116 competent cells are now available lyophilized. LyoComp GT116 are easy to handle and more stable than standard competent cells. They are shipped at room temperature eliminating costly dry ice shipping. They can be stored at 4ºC for short term storage or at –20ºC for long term storage. Their competency will allow you to successfully clone into psiRNA plasmids. Sequencing Primers Hairpin structures are known to be difficult to sequence. Therefore, InvivoGen designed several primers located 5’ or 3’ to the insert and selected a pair, OL381 and OL178, that allows the full reading of the insert sequence by using conventional sequencing methods. E. coli Fast-Media® XGal To facilitate the selection of recombinant clones, pouches of Fast–Media® Zeo Xgal or Kan Xgal are provided for the preparation in less than 5 minutes of high quality E. coli medium. Fast–Media® is a ready-to-use microwaveable LB based medium powder that already contains the antibiotic, IPTG and X-Gal at the right concentration. 1. Connelly JC. et al., 1998. The SbcCD nuclease of Escherichia coli is a structural maintenance of chromosomes (SMC) family protein that cleaves hairpin DNA. Proc. Natl. Acad. Sci. USA, 95:7969-7974   Contents and Storage E. coli GT116 is provided either as a lyophilized strain on paper disk or as chemically competent cells. Sequencing Primers are provided as 10 µg each of OL381 and OL178 lyophilized oligonucleotides. E. coli Fast-Media® XGal is provided in a 20-pouch unit. Each pouch enables the preparation of 8-10 agar plates. Each psiRNA kit contains 20 µg of psiRNA plasmid, 1 disk of E. coli GT116 strain, 20 µg of sequencing primers, 4 pouches of the appropriate Fast-Media® XGal. Products are shipped at room temperature. Fast-Media® pouches should be stored at room temperature. Store all other components at –20°C.

Components of the psiRNA™ System are available individually or in a kit. Choose from five different psiRNA kits: psiRNAzeo G2 kit psiRNAneo G2 kit psiRNAblasti G2 kit psiRNAhygro G2 kit psiRNAGFPzeo G2 kit Product Quantity Catalog code psiRNA G2 Kits – Zeo – Neo – Blasti – Hygro – GFPzeo - - - - - k-psirna-zeog2 k-psirna-neog2 k-psirna-blastig2 k-psirna-hygrog2 k-psirna-gfpzeog2 psiRNA Vectors 20 µg see psiRNA Vectors E. coli GT116  – Lyophilized  – LyoComp 1 paper disk 5x 0.2ml (lyophilized) bac–gt116 lyo-116-21 Sequencing primer 2 x 10 µg ol–381-178 FastMedia® XGal – Zeo – Kan – Blasti – Hygro 20 pouches 20 pouches 20 pouches 20 pouches fas-zn-x fas-kn-x fas-bl-x fas-hg-x