BLOCK-iT™ Lentiviral Pol II miR RNAi Expression System (Invitrogen™)

The BLOCK-iT™ Lentiviral Pol II miR RNAi Expression System combines
Invitrogen's BLOCK-iT™ Pol II miR RNAi and ViraPower™ Lentiviral technologies
to facilitate creation of a replication-incompetent lentivirus that delivers a
microRNA (miRNA) sequence of interest to dividing or non-dividing mammalian
cells for RNA interference (RNAi) analysis thus broadening the
potential RNAi applications beyond those of other traditional retroviral
systems (Naldini, 1998).

The BLOCK-iT™ Lentiviral Pol II miR RNAi Expression System includes:
o A BLOCK-iT™ Pol II miR RNAi Expression Vector Kit for production of an
expression clone containing a double-stranded oligonucleotide encoding a pre-miRNA sequence for expression in mammalian cells. The BLOCK-iT™ Pol II miR RNAi Expression Vector (pcDNA™6.2-GW/miR) provides a rapid and efficient way to clone ds oligo duplexes encoding a desired miRNA target sequence into a vector containing a Pol II promoter (CMV) for use in RNAi analysis. The BLOCK-iT™ Pol II miR RNAi Expression Vector is specifically designed to
allow expression of miRNA sequences and contain specific miR flanking
sequences that allow proper processing of the miRNA.
o The pDONR™221 vector to transfer the premiRNA
expression cassette into the lentiviral expression plasmid using Gateway™ Technology. The Gateway™ Technology is a universal cloning method that takes advantage of the site-specific recombination properties of bacteriophage lambda (Landy, 1989) to provide a rapid and highly efficient way to move your DNA sequence of interest into multiple vector systems.
o A pLenti6/V5-DEST destination vector into which the pre-miRNA cassette
from the expression clone is transferred using Gateway™ Technology.This expression plasmid contains elements that allow packaging of the construct into virions and the Blasticidin resistance marker for selection of stably transduced cell lines.
o Gateway™ BP and LR Clonase™ II Enzyme Mixes that facilitate the transfer
of the pre-miRNA expression cassette from the expression vector into the
pLenti6/V5-DEST destination vector.
o Components of the ViraPower™ Lentiviral System for production of a
replication-incompetent lentivirus that stably expresses the miRNA of
interest in both dividing and non-dividing mammalian cells. The ViraPower™ Lentiviral Technology facilitates highly efficient, in vitro or invivo delivery of a target gene or RNA to dividing and non-dividing mammalian cells using a replication-incompetent lentivirus. Based on the lentikat™ systemdeveloped by Cell Genesys (Dull et al., 1998), the ViraPower™ LentiviralTechnology possesses features which enhance its biosafety while allowing highlevel expression in a wider range of cell types than traditional retroviral systems.