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A versatile approach to multiple gene RNA interference using microRNA-based short hairpin RNAs

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Author(s): Zhu Xiaocui | Santat Leah | Chang Mi | Liu Jamie | Zavzavadjian Joelle | Wall Estelle | Kivork Christine | Simon Melvin | Fraser Iain

Journal: BMC Molecular Biology
ISSN 1471-2199

Volume: 8;
Issue: 1;
Start page: 98;
Date: 2007;
Original page

ABSTRACT
Abstract Background Effective and stable knockdown of multiple gene targets by RNA interference is often necessary to overcome isoform redundancy, but it remains a technical challenge when working with intractable cell systems. Results We have developed a flexible platform using RNA polymerase II promoter-driven expression of microRNA-like short hairpin RNAs which permits robust depletion of multiple target genes from a single transcript. Recombination-based subcloning permits expression of multi-shRNA transcripts from a comprehensive range of plasmid or viral vectors. Retroviral delivery of transcripts targeting isoforms of cAMP-dependent protein kinase in the RAW264.7 murine macrophage cell line emphasizes the utility of this approach and provides insight to cAMP-dependent transcription. Conclusion We demonstrate functional consequences of depleting multiple endogenous target genes using miR-shRNAs, and highlight the versatility of the described vector platform for multiple target gene knockdown in mammalian cells.
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