Ez chip chromatin immunoprecipitation kit for cell line samples

We performed chromatin immunoprecipitation (ChIP) using the EZ ChIP™Chromatin Immunoprecipitation Kit for cell line samples (Millipore, Bedford, MA). Briefly, we sonicated the crosslinked chromatin DNA into 200-to 500-bp fragments. Normal mouse IgG was used as the negative control. The primer sequences are listed in Table 3. The antibodies for the ChIP assays were obtained from Millipore. Quantification of the immunoprecipitated DNA was performed using qPCR with SYBR Green Mix (Takara). The ChIP data were calculated as a percentage relative to the input DNA using the equation 2[Input Ct-Target Ct] × 0.1 × 100.

The ChIP experiments were performed using an EZ ChIP™ Chromatin Immunoprecipitation kit for cell line samples (Millipore, Billerica, MA, uSA) according to the manufacturer's instructions. Briefly, the crosslinked chromatin DNA were sonicated into 200-1000 bp fragments and fixed with 1% formaldehyde. Then immunoprecipitation followed using an anti-DNMT1 antibody and normal mouse IgG as the negative control. The primers used for the amplification of RIZ1 promoter DNA fragments are listed in Table I.
Statistical analysis. The independent Student's t-test was used to compare the results expressed as mean ± SD and Chi-square (χ 2 ) test for categorical variables between any two pre-selected groups. A P-value <0.05 was considered to indicate a statistically significant difference.

Chromatin immunoprecipitation (ChIP) experiments were performed using an EZ ChIP™ Chromatin Immunoprecipitation Kit for cell line samples (Millipore, Billerica, MA) according to the manufacturer's instructions. Briefly, the crosslinked chromatin DNA was sonicated into 200 to 1000 bp fragments and then fixed with 1% formaldehyde. Immunoprecipitation was performed using an anti-PITX1 antibody or normal mouse IgG as the negative control. All standard protocols for sequence preparation, sequencing, and quality control were provided by Illumina (San Diego, CA). In short, DNA recovered from a conventional ChIP procedure was quantified using a QuantiFluor fluorometer (Promega, Madison, WI). DNA integrity was verified using the Agilent Bioanalyzer 2100 (Agilent, Palo Alto, CA). The DNA was then processed, including end-repair, adaptor ligation, and size selection, using an Illumina sample prep kit following the manufacturer's instructions (Illumina). Final DNA libraries were validated and sequenced at 75 bp per sequence read using an Illumina GAIIx sequencer at a depth of approximately 30 million sequences per sample.