title: GSM3703979: LeLab_ChIPseq_TLE3000_pUT18C_1xFLAG_Dechloromonas_aromatica_ParB; Escherichia coli; ChIP-Seq
library_construction_protocol: For ChIP-seq, DNA was isolated using Qiagen PCR purification columns. For B1H-seq, plasmid DNA was isolated using Qiagen MiniPrep columns. Detailed protocols are listed in the Supplementary Materials For ChIP-seq libraries, a standard library construction procedure (using NEBNext® Ultra™ II DNA Library Prep Kit for Illumina sequencing) was employed. For B1H-seq, a similar strategy to amplicon-seq was used to construct libraries for Illumina sequencing. Briefly,Illumina Truseq-compatible libraries were constructed from pre- and post-selection plasmid libraries via two rounds of PCR.  PCR round 1:  Primer 4nns_R (10 µM): 2.5 µL  Mixture in equimolar amount of primers 4nns_offset_0_F; 4nns_offset_1_F; 4nns_offset_2_F; 4nns_offset_3_F; 4nns_offset_4_F (10 µM): 2.5 µL. A mixture of forward primers were used to stagger reads across the amplicon to improve the distribution of base calls at each position during the initial rounds of Illumina sequencing.  dNTP (10mM): 1 µL  DMSO: 1.5 µL  Plasmid template (pre- or post-selection): 1 µL of 500 ng/µL  Phusion polymerase: 0.5 µL  5x HF buffer: 10 µL  Water: 31 µL  PCR program: 98oC for 30s, (98oC for 10s, 56oC for 20s, 72oC for 10s) x 20 cycles, 72oC for 5min.  PCR products were gel-purified, quantified by Qubit hsDNA quantification kit (ThermoFisher), and used as template in the second PCR.  PCR round 2:  NEBNext Index primer (NEB): 2.5 µL  NEBNext universal primer (NEB): 2.5 µL  dNTP (10mM): 1 µL  DMSO: 1.5 µL  Template: 5 µL of gel-purified DNA from PCR round 1  5x HF buffer: 10 µL  Phusion polymerase: 0.5 µL  Water: 27 µL  PCR program: 98oC for 30s, (98oC for 10s, 54oC for 20s, 72oC for 10s) x 12 cycles, 72oC for 5min.  PCR products were gel-purified, quantified by Qubit hsDNA quantification kit (ThermoFisher), and were sequenced on the Illumina HiSeq 2500 (single-end, 150 bp read length, 15% spike-in phiX DNA) at the Tufts University Genomics facility.