Detecting expressed genes using CAGE.

Cap analysis of gene expression (CAGE) provides accurate high-throughput measurement of RNA expression. By the large-scale analysis of 5' end of transcripts using CAGE method, it enables not only determination of the transcription start site but also prediction of promoter region. Here we provide a protocol for the construction of no-amplification non-tagging CAGE libraries for Illumina next-generation sequencers (nAnT-iCAGE). We have excluded the commonly used PCR amplification and cleavage of restriction enzyme to eliminate any potential biases. As a result, we achieved less biased simple preparation process.