Merck

Lentivirus-delivered stable gene silencing by RNAi in primary cells.

RNA (New York, N.Y.) (2003-03-22)
Sheila A Stewart, Derek M Dykxhoorn, Deborah Palliser, Hana Mizuno, Evan Y Yu, Dong Sung An, David M Sabatini, Irvin S Y Chen, William C Hahn, Phillip A Sharp, Robert A Weinberg, Carl D Novina
ABSTRACT

Genome-wide genetic approaches have proven useful for examining pathways of biological significance in model organisms such as Saccharomyces cerevisiae, Drosophila melanogastor, and Caenorhabditis elegans, but similar techniques have proven difficult to apply to mammalian systems. Although manipulation of the murine genome has led to identification of genes and their function, this approach is laborious, expensive, and often leads to lethal phenotypes. RNA interference (RNAi) is an evolutionarily conserved process of gene silencing that has become a powerful tool for investigating gene function by reverse genetics. Here we describe the delivery of cassettes expressing hairpin RNA targeting green fluorescent protein (GFP) using Moloney leukemia virus-based and lentivirus-based retroviral vectors. Both transformed cell lines and primary dendritic cells, normally refractory to transfection-based gene transfer, demonstrated stable silencing of targeted genes, including the tumor suppressor gene TP53 in normal human fibroblasts. This report demonstrates that both Moloney leukemia virus and lentivirus vector-mediated expression of RNAi can achieve effective, stable gene silencing in diverse biological systems and will assist in elucidating gene functions in numerous cell types including primary cells.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Bacterial Glycerol Stock, Tumor Suppressors
Sigma-Aldrich
MISSION® shRNA Mouse Gene Family Set, Bacterial Glycerol Stock, Tyrosine Kinases
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Bacterial Glycerol Stock, Ion Channels
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Bacterial Glycerol Stock, Cytokines and Chemokines
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Bacterial Glycerol Stock, Kinases
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Cytokines and Chemokines
Sigma-Aldrich
MISSION® Lentiviral Packaging Mix, Production of replication incompetent viral particles
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Ion Channels
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, G-Protein Coupled Receptors
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Cytokine and Chemokine Receptors
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Apoptosis Pathway
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Nuclear Hormone Receptors
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Tumor Suppressors
Sigma-Aldrich
MISSION® shRNA Human Gene Family Set, Lentiviral Particles, Kinases
Sigma-Aldrich
MISSION® pLKO.1-puro TurboGFP shRNA Control Plasmid DNA, shRNA sequence targeting tGFP