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. 1988 Jul 11;16(13):5999–6014. doi: 10.1093/nar/16.13.5999

Use of RNase H and primer extension to analyze RNA splicing.

S H Erster 1, L A Finn 1, D A Frendewey 1, D M Helfman 1
PMCID: PMC336843  PMID: 2840638

Abstract

A new method for the characterization of pre-mRNA splicing products is presented. In this method RNA molecules are hybridized to an oligodeoxynucleotide complementary to exon sequences upstream of a given 5' splice site, and the RNA strands of the resulting RNA:DNA hybrids are cleaved by RNase H. The cleaved RNAs are then subjected to primer extension using a 32P-labelled primer complementary to exon sequences downstream of an appropriate 3' splice site. Since the primer extension products all terminate at the site of RNase H cleavage, their lengths are indicative of the splice sites utilized. The method simplifies the study of the processing of complex pre-mRNAs by allowing the splicing events between any two exons to be analyzed. We have used this approach to characterize the RNAs generated by expression of the rat tropomyosin 1 (Tm 1) gene in various rat tissues and in cultured cells after transient transfection. The results demonstrate that this method is suitable for the analysis of alternative RNA processing in vivo.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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