Affinity-seq of mouse and human ZFY proteins and mouse CTCF

This experiment determines all possible genome-wide binding sites of the proteins of interest The proteins of interest were expressed in E. coli as fused proteins with an N-terminal 6HisHALO tag and partially purified on SP-sepharose. Mouse spleen genomic DNA was sheard to an average of 200 bp fragments and used as binding target. The partially purified protein and the sheared genom DNA were allowed to bind in solution and the complexes were then isolated on Ni2+ beads preincubated with expressed 6HihHALO tag only to reduce the background. The eluted DNA was used to prepare NextGen libraries which were sequenced on Illumina HiSeq 2000.