GSM8014137: GmCOT_GLYMA.08G227700-ChIP_br2; Glycine max; ChIP-Seq
Chromatin isolation and immunoprecipitation were performed as described in Gendrel AV, Lippman Z, Martienssen R, Colot V. Nat Methods. 2005;2:213–218, except that the isolated nuclei were resuspended in shearing buffer (10mM Tris pH8, 1mM EDTA, 0.1% SDS, 1mM PMSF), sonicated to achieve chromatin fragments ranging from 100 to 500 bp, and Triton X-100 and NaCl were added to chromatin to final concentrations of 1% and 150mM, respectively. Nuclear extracts were either set aside for whole genome extracts (input control) or incubated overnight with 3 ug of an antibody raised against the peptide FSTQPGDHETESDVN corresponding to the amino acids 352-366 of GLYMA.08G227700.  Crosslinks were reversed and protein digested as described in Dahl and Collas, Nat Protoc. 2008;3(6):1032-45, extracted with phenol:chloroform and precipitated using standard procedures. ChIP and Input libraries were generated using Nugen Ovation Ultralow System V2, following the manufaturer's instructions and enriched by PCR using 15 cycles.  Libraries were size-selected by electrophoresis and purified using Qiagen MinElute gel purification kit