Epiquick tissue chromatin immunoprecipitation kit

Chromatin immunoprecipitation (ChIP) assays were carried out as previously described (Simon et al., 2012) . Briefly, ChIP assays were performed on cortical tissue collected from ten wild-type embryos at E14.5 employing a specific mouse monoclonal antibody recognizing Bcl11a (Abcam Cat# ab19489, RRID: AB_2063996) and using the EpiQuick Tissue Chromatin Immunoprecipitation Kit (Epigentek), including mouse IgG as a non-specific control. The precipitated DNA was analyzed by quantitative real-time PCR using oligonucleotides recognizing a 1.1-kb region in the second intron of Sema3c: 5 0 -GCGCCAGAGAACCTGACA-3 0 and 5 0 -CTGCTGCTGTGGCTTAGG-3 0 . As a negative control, the Hprt promoter region was used, which was recognized by the following oligonucleotides: 5 0 -CTGCCTCTGCCTCCTAAATG-3 0 and 5 0 -TGTCGTCTCCCAGAGGATTC-3 0 . ChIP quantitative real-time PCR data were analyzed by the comparative C T method determining the fold enrichment of the immunoprecipitated DNA by the specific antibody versus IgG using the input DNA as reference. C T values > 35 were disregarded. Every ChIP experiment was repeated at least three times.