1. A versatile approach to multiple gene RNA interference using microRNA-based short hairpin RNAs.
- Author
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Xiaocui Zhu, Santat, Leah A., Mi Sook Chang, Liu, Jamie, Zavzavadjian, Joelle R., Wall, Estelle A., Kivork, Christine, Simon, Melvin I., and Fraser, Iain D. C.
- Subjects
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RNA , *GENES , *CLONING , *RETROVIRUS diseases , *PROTEIN kinases , *MAMMALS - 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. [ABSTRACT FROM AUTHOR]
- Published
- 2007
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