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Nucleic Acids Research, 2003, Vol. 31, No. 6 e24
© 2003 Oxford University Press

Small amplified RNA-SAGE: an alternative approach to study transcriptome from limiting amount of mRNA

Catheline Vilain1, Frederick Libert1, David Venet1, Sabine Costagliola1 and Gilbert Vassart1,2

1 Institute of Interdisciplinary Research (IRIBHM) and 2 Department of Medical Genetics of the Erasme Hospital, Université Libre de Bruxelles, 808 route de Lennik, B-1070 Bruxelles, Belgium

*To whom correspondence should be addressed. Tel: +32 2 555 41 97; Fax: +32 2 555 46 55; Email: cavilain{at}ulb.ac.be

Serial analysis of gene expression (SAGE) is a widely used and powerful technique to characterize and compare transcriptomes. Although several modifications have been proposed to the initial protocol with the aim of reducing the amount of starting material, unless additional PCR steps are added, the technique is still limited by the need for at least 1 µg of total RNA. As extra PCR amplification might introduce representation biases, current SAGE protocols are not fully suitable for the study of small, microdissected tissue samples. We propose here an alternative method involving the linear amplification of small mRNA fragments containing the SAGE tags. The procedure allows preparation of libraries of over 100 000 tags from as few as 2500 cells. A satisfactory correlation was observed between a microSAGE library made from 5 µg of total thyroid RNA, and a library prepared from 50 ng of the same RNA preparation according to the present protocol.


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