Effect of depletion of Hp1α (Cbx5) and Hp1β (Cbx1) on gene expression of mouse embryonic stem cells (mESCs)

To investigate the function of HP1α and HP1β proteins in the regulation of heterochromatin in mESCs, we isolated blastocysts from matings of mice homozygous for Hp1βF/F; Hp1αF/F in a single well of a 96-well plate in ESC medium. After 2 days, when blastocysts hatched, they were trypsinized and cells were seeded in the same well for other 2 days. Expanded mESC clones were further tested for their genotype by PCR. A single clone of Hp1βF/F; Hp1αF/F was transfected with either pCAG-Cre-GFP or pCAG-GFP as a control. GFP positive cells were sorted by flow cytometry and seeded into 10 cm plates. Individual clones were isolated and expanded. Deletion of Hp1β or Hp1α floxed alleles in pCAG-Cre-GFP transfected clones was tested by Western Blotting with appropriate antibodies. We then performed gene expression profiling analysis using data obtained from RNA-seq from 2 different clones with deleted Hp1α and Hp1β and 1 control clone.