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Published online 2 January 2004

Nucleic Acids Research, 2004, Vol. 32, No. 1 e6
© 2004 Oxford University Press

Improved full-length cDNA production based on RNA tagging by T4 DNA ligase

Christian Clepet*, Isabelle Le Clainche and Michel Caboche

Unité de Recherches en Génomique Végétale, INRA/CNRS, 2 Rue Gaston-Crémieux, F-91057 Evry Cedex, France

*To whom correspondence should be addressed. Tel: +33 160874512; Fax: +33 160874510; Email: clepet{at}evry.inra.fr

Second-strand cDNA priming is a central problem for full-length characterization of transcripts. A new strategy using bacteriophage T4 DNA ligase and partially degenerate adapters is proposed for grafting a sequence tag to the end of polyribonucleotides. Based on this RNA tagging system and previously described protocols, a new method for full-length cDNA production has been implemented. Validation of the method is shown in Arabidopsis thaliana by the construction of a full-length cDNA library and the analysis of 154 clones and by 5'-RACE–PCR run on a documented experimental system.


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