Feed-forward regulation of a cell fate determinant by an RNA-binding protein generates asymmetry in yeast
Khd1-TAP was purified from 1L of cells grown to OD600 2.5 and UV-crosslinked three times at 400 mJ/cm2. Purification using calmodulin sepharose was followed by binding to magnetic IgG beads. The CLIP protocol was then followed as previously described (ULE et al. 2005). The resulting cDNA was amplified using PCR with oligonucleotides containing sequences for hybridization to the Illumina flow cell.