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Biochem Biophys Res Commun. 2007 Jun 29;358(2):521-7. Epub 2007 May 7.

Achieving targeted and quantifiable alteration of mRNA splicing with Morpholino oligos.

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  • 1Gene Tools LLC, Philomath, OR 97370, USA. pmorcos@gene.tools.com

Abstract

This work represents the first guide for using steric-block antisense oligos as tools for effective and targeted modification of RNA splicing. Comparison of several steric-block oligo types shows the properties of Morpholinos provide significant advantages over other potential splice-blocking oligos. The procedures and complications of designing effective splice-blocking Morpholino oligos are described. The design process requires complete pre-mRNA sequence for defining suitable targets, which usually generate specific predictable messengers. To validate the targeting procedure, the level and nature of transcript alteration is characterized by RT-PCR analysis of splice modification in a beta-globin splice model system. An oligo-walking study reveals that while U1 and U2 small nuclear RiboNucleoProtein (snRNP) binding sites are the most effective targets for blocking splicing, inclusion of these sites is not required to achieve effective splice modifications. The most effective targeting strategy employs simultaneously blocking snRNP binding sites and splice-junctions. The work presented here continues to be the basis for most of the successful Morpholino oligos designed for the worldwide research community to block RNA splicing.

PMID:
17493584
DOI:
10.1016/j.bbrc.2007.04.172
[PubMed - indexed for MEDLINE]
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