期刊
RNA
卷 9, 期 4, 页码 493-501出版社
COLD SPRING HARBOR LAB PRESS, PUBLICATIONS DEPT
DOI: 10.1261/rna.2192803
关键词
retrovirus; hairpin RNA; siRNA; knockdown; dendritic cells
资金
- NCI NIH HHS [F32 CA093033, KO1CA94223, K01 CA094223, P30 CA014051, P30-CA14051, F32 CA93033-01, P01-CA42063, R01 CA078461, R01CA78461-04, P01 CA042063] Funding Source: Medline
- NIAID NIH HHS [F32 AI010523, R01-AI32486, F32 AI10523] Funding Source: Medline
- NIGMS NIH HHS [R37 GM034277, R37-GM34277] Funding Source: Medline
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.
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