title: GSM8801440: B2-1_mCheHI_CitHI; Homo sapiens; OTHER
library_construction_protocol: Genomic DNA was extracted with a DNEasy Blood and Tissue Kit (QIAgen) following manufacturer's protocol for each bin extracted from the sorting experiment (at 1e5 cells). DNA was eluted in EB and sequences were amplified by PCR. A two-step PCR process was used to append Illumina adapters as overhangs. 6x reactions were prepared with 1ug of genomic DNA, 5uL of each 10uM primer, and 25uL of NEBnext 2x Master Mix (NEB) was used for the first reaction. PCR amplification parameters were as follows: 3 minutes at 98˚C, then 21x cycles of 98˚C for 10 sec, 68˚C for 30 sec, 72˚C for 30 sec, and a final step of 72˚C for 2 minutes. Resulting DNA was gel extracted using a QIAgen Gel Extraction Kit (QIAgen). A second PCR was performed with the Illumina indexed primers with the following PCR amplification parameters: 3 minutes at 98˚C, then 11x cycles of 98˚C for 10 sec, 68˚C for 30 sec, 72˚C for 30 sec, and a final step of 72˚C for 2 minutes. Resulting DNA was again gel extracted using a QIAgen Gel Extraction Kit (QIAgen). Libraries were then quantified with a Qubit HS dsDNA High Sensitivity Kit (Thermo Fisher), pooled with 15% PhiX control (Illumina), and sequenced on an Illumina HiSeq 3000.