title: GSM4551298: ATAC_CD34_BCLKO32_rep1; Homo sapiens; ATAC-seq
library_construction_protocol: Omni-ATAC was performed with 50,000 fresh cells or frozen pellets. Cells were first permeabilized with 50 uL RSB buffer (10 mM Tris-HCl pH 7.4, 10 mM NaCl, 3 mM MgCl2) with 0.1% NP40, 0.1 Tween-20 and 0.01% Digitonin for 3 minutes. 1 mL of cold RSB containing 0.1% Tween-20 was added before cells were pelleted again. The pellets were then resuspended with 50 uL transposition mixture (25uL 2xTD buffer from ATACseq kit, 2.5 uL transposase, 16.5 uL PBS, 0.5 ul 1% digitonin, and 0.5 ul 10% Tween-20), and reaction was carried out at 37 °C for 60 minutes. Genomic DNA was extracted with Qiagen minElute PCR purification kit according to manufacturer's manual. Total DNA was used as template for amplification, mixed with 15 uL of 2x Ultra II Q5 mix, 1 uL of 10 uM i7 primer (AATGATACGGCGACCACCGAGATCTACAC-TCGTCGGCAGCGTCAGATGTG) and 1 uL of 10 uM i5 primer (CAAGCAGAAGACGGCATACGAGATNNNNNNNNGTCTCGTGGGCTCGGAGATGT) where N represent barcodes. PCR amplification was performed as follow: 65°C 5 min, 98°C 30 s, 98°C 10 s, 63°C 30 s, repeat step 3-4 for 9 times, and 63°C 5min. PCR products were purified with AMPure beads and quantified using Qubit and Tapestation before pooling and sequencing. Total tagmentated genomic DNAwas  mixed with 15 uL of 2x Ultra II Q5 mix, 1 uL of 10 uM i7 primer (AATGATACGGCGACCACCGAGATCTACAC-TCGTCGGCAGCGTCAGATGTG) and 1 uL of 10 uM i5 primer (CAAGCAGAAGACGGCATACGAGATNNNNNNNNGTCTCGTGGGCTCGGAGATGT) where N represent barcodes. PCR amplification was performed as follow: 65°C 5 min, 98°C 30 s, 98°C 10 s, 63°C 30 s, repeat step 3-4 for 9 times, and 63°C 5min. PCR products were purified with AMPure beads and quantified using Qubit and Tapestation before pooling and sequencing. Tn-seq
attributes.tag: GEO Accession
attributes.value: GSM4551298