Nucleic Acids Research, Vol 27, Issue 5 1300-1307, Copyright © 1999 by Oxford University Press
NA Datson, J van der Perk-de Jong, MP van den Berg, ER de Kloet and E Vreugdenhil
Serial Analysis of Gene Expression (SAGE) is a powerful expression
profiling method, allowing the analysis of the expression of thousands of
transcripts simultaneously. A disadvantage of the method, however, is the
relatively high amount of input RNA required. Consequently, SAGE cannot be
used for the generation of expression profiles when RNA is limited, i.e. in
small biological samples such as tissue biopsies or microdissected
material. Here we describe a modification of SAGE, named microSAGE, which
requires 500- to 5000-fold less starting material. Compared with SAGE,
microSAGE is simplified due to incorporation of a 'single-tube' procedure
for all steps from RNA isolation to tag release. Furthermore, a limited
number of additional PCR cycles are performed. Using microSAGE gene
expression profiles can be obtained from minute quantities of tissue such
as a single hippocampal punch from a rat brain slice of 325 micrometers
thickness, estimated to contain, at most, 10(5) cells. This method opens up
a multitude of new possibilities for the application of SAGE, for example
the characterization of expression profiles in tissue biopsies, tumor
metastases or in other cases where tissue is scarce and the generation of
region-specific expression profiles of complex heterogeneous tissues.
ARTICLES
MicroSAGE: a modified procedure for serial analysis of gene expression in limited amounts of tissue
Division of Medical Pharmacology, Leiden/Amsterdam Center for Drug Research, Leiden University, PO Box 9503, 2300 RA Leiden, The Netherlands. datson_n@lacdr.leidenuniv.nl
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