title: GSM5915587: HSC on AAVS1 with IDLV Mouse 1; Homo sapiens; OTHER
library_construction_protocol: gDNA was isolated with QIAamp DNA Micro Kit (QIAGEN) according to the manufacturer's instructions. PCR amplicons for individual samples were generated by nested PCR using primers listed in the related manuscript and starting from >50-100 ng of purified gDNA. The first PCR step was performed with GoTaq G2 DNA Polymerase (Promega) according to manufacturer instruction using the following amplification protocol: 95°C x 5'min, (95°C x 0.5 min, 60°C x 0.5 min, 72°C x 0.2 5min) x 20 cycles, 72°C x 5 min. The second PCR step was performed with GoTaq G2 DNA Polymerase (Promega) according to manufacturer instruction using 5μl of the first-step PCR product and the following amplification protocol: 95°C x 5min, (95°C x 0.5 min, 60°C x 0.5 min, 72°C x 0.3 min) x 20 cycles, 72°C x 5 min. Second-step PCR primers were endowed with tails containing P5/P7 sequences, i5/i7 Illumina tags to allow multiplexed sequencing and R1/R2 primer binding sites (Supplementary Table 5). PCR amplicons were separately purified performing double-side selection with AmpPure XP beads (Beckman Coulter) or QIAquick PCR Purification Kit (QIAGEN). Amplicons concentration and quality was assessed by QuantiFluor ONE dsDNA system and 4200 Tapestation System (Agilent). Amplicons from up to 36 differently tagged samples were multiplexed at equimolar ratios and run