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Serial analysis of gene expression

Nature Protocols volume 1, pages 1743–1760 (2006)Cite this article

Abstract

Serial analysis of gene expression (SAGE) is a method used to obtain comprehensive, unbiased and quantitative gene-expression profiles. Its major advantage over arrays is that it does not require a priori knowledge of the genes to be analyzed and reflects absolute mRNA levels. Since the original SAGE protocol was developed in a short-tag (10-bp) format, several modifications have been made to produce longer SAGE tags for more precise gene identification and to decrease the amount of starting material necessary. Several SAGE-like methods have also been developed for the genome-wide analysis of DNA copy-number changes and methylation patterns, chromatin structure and transcription factor targets. In this protocol, we describe the 17-bp longSAGE method for transcriptome profiling optimized for a small amount of starting material. The generation of such libraries can be completed in 7–10 d, whereas sequencing and data analysis require an additional 2–3 wk.

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Figure 1: Schematic outline of the SAGE protocol.
Figure 2: Image of gel with ditag PCR products.
Figure 3: Image of gel of ditag fragments and linkers released by NlaIII digest.
Figure 4: Image of gel of ditag concatemer formation.

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  1. Department of Medical Oncology, Dana-Farber Cancer Institute, Harvard Medical School, 44 Binney Street, D740C, Boston, 02115, Massachusetts, USA

    Min Hu & Kornelia Polyak

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Correspondence to Kornelia Polyak.

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Competing interests

MH and KP are inventors of a patent application submitted on the MSDK (Methylation Specific Digital Karyotyping) technology and DNA methylation markers identified with it. KP receives research support from and is a consultant to Novartis Pharmaceuticals Inc.

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Hu, M., Polyak, K. Serial analysis of gene expression. Nat Protoc 1, 1743–1760 (2006). https://doi.org/10.1038/nprot.2006.269

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