SAGE analysis from 1 microg of total RNA

Serial analysis of gene expression (SAGE) is a powerful transcription-profiling method that allows simultaneous expression analysis of thousands of transcripts, provides absolute digital readout of the expression level, and identifies new genes. A disadvantage of SAGE is the relatively high amount of input RNA required. Consequently, several techniques have been developed to overcome this limitation, so that SAGE can be applied to very limited amounts of starting material, e.g., small biological samples such as tissue biopsies or microdissected materials. Here we describe a modified version of the original microSAGE protocol, which requires only 1 to 2 microg total RNA. This method avoids PCR amplification of cDNA and reamplification of ditags, procedures that potentially compromise the quantitative nature of this technique.