ABSTRACT
This chapter presents an established Cap Analysis of Gene Expression (CAGE) protocol to work with the sequencer GS20/GSFLX system. CAGE tags are derived from the 5’ end of ribonucleic acids expressed in the cells and tissues. CAGE allows high-throughout gene expression analysis and the simultaneous profiling of transcriptional start site usage and identification of core promoters. Cethyl trimethyl ammonium bromide is a cationic detergent that with controlled NaCl concentration will specifically precipitate long nucleic acids by creating specific complexes left in solution with other molecules like proteins, short oligonucleotides and small molecules. By pooling, all the CAGE tags are subjected at the same Polymerase Chain Reaction (PCR) condition in the same reaction, minimizing differences among samples due to PCR. Perform the purification as suggested in the protocol attached to the Qiagen MinElute kit, following by deoxyribonucleic acids pooling and concentration by ethanol purification.
