title: GSM7675479: ABE8.20-m at AAVS1 locus on mPB CD34 edited at day 3 collected at day 7 - rep4; Homo sapiens; OTHER
library_construction_protocol: gDNA was isolated with QIAamp DNA Micro Kit (QIAGEN) according to the manufacturer's instructions. Custom library construction protocol. PCR amplicons for each individual sample were generated by nested PCR starting from >50-100 ng of purified gDNA.The first PCR step was performed using the following amplification protocol: 95°C x 10', (95°C x 30'', 59°C x 30'', 72°C x 20'') x 20 cycles, 72°C x 10'. The second PCR step was performed using the same reagents of the first step, 5 μl of the PCR product and the following amplification protocol: 95°C x 10', (95°C x 30'', 59°C x 30'', 72°C x 20'') x 20 cycles, 72°C x 10'. Primers used for the second PCR step contained P5/P7 sequences, i5/i7 Illumina tags to allow multiplexed sequencing and R1/R2 primer binding sites. The PCR amplicon from each sample was separately purified by QIAquick PCR Purification Kit (QIAGEN). Concentration and quality of amplicons were assessed by QuantiFluor ONE dsDNA system and 4200 Tapestation System (Agilent). Amplicons from up to 25 differently tagged samples were multiplexed at equimolar ratios.