Re-stimulated control or Eomes-overexpressing OT-I cells were harvested, fixed with 1% formaldehyde for 10min at room temperature and stopped by adding glycine to 125mM. ChIP experiment was done following instructions of Active Motif's ChIP assay kit (53035) with slight modifications, in which 10 cycles of sonication (Bioruptor) were added after enzyme digestion, and dynabeads protein G (Life Technologies) were used for immunoprecipitations. 5μg anti-TBR2/Eomes antibodies (ab23345, abcam) were used for each reaction. The precipitated DNA was sent to BGI genomics for library construction.