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Methods

A tale of two templates: Automatically resolving double traces has many applications, including efficient PCR-based elucidation of alternative splices

¹ Laboratory for Computational Genomics and Department of Computer Science, Washington University, St. Louis, Missouri 63130, USA; ² Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, Connecticut 06620-8103, USA; ³ Department of Biology, Washington University, St. Louis, Missouri 63130, USA

Trace Recalling is a novel method for deconvoluting double traces that result from simultaneously sequencing two DNA templates. Trace Recalling identifies up to two bases at each position of such a trace. The resulting ambiguity sequence is aligned to the genome, identifying one template sequence. A second template sequence is then inferred from this alignment. This technique makes possible many exciting biological applications. Here we present two such applications, alternate splice finding and elucidation of multiple insertion sites in a random insertional mutagenesis library. Our results demonstrate that RT–PCR followed by Trace Recalling is a more efficient and cost effective way to find alternate splices than traditional methods. We also present a method for mapping double-insertion events in a random insertional-mutagenesis library.

E-mail brent{at}cse.wustl.edu; fax (314) 935-7302.

⁴ These two authors contributed equally to this work.

[Supplemental material is available online at www.genome.org. Alternate splice forms discovered during this work have been deposited in GenBank under accession nos. EB71062–EB710342.]

Article published online before print. Article and publication date are at http://www.genome.org/cgi/doi/10.1101/gr.5661407

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