GSM4649357: WT; Homo sapiens; OTHER
Cells were lysed in 10 mM Tris-HCl, pH 8.0, 100 mM EDTA, 0.5% SDS, 200 μg/ml RNase A and incubated at 37 °C for 1 hour followed by the addition of 1mg/ml of proteinase K for an overnight incubation at 50 °C. The resulting lysates were extracted three times with phenol solution equilibrated with 10 mM Tris-HCl, pH 8.0, 1 mM EDTA (Sigma P4557) mixed with chloroform (1:1 v/v) using tubes with phase lock gel (Tiangen WM5-2302831). The aqueous phase was then mixed with 0.2 volume of 10 M ammonium acetate and 1 volume of isopropanol, resulting the immediate formation of cloudy DNA precipitates. Precipitated DNA was then transferred by a pipet tip to a tube containing 75% (v/v) ethanol. This process was repeated twice to ensure complete removal of residual organic solvents. Afterwards, genomic DNA was dissolved in 1 mM Tris-HCl, pH 8.0, 0.1 mM EDTA by incubation at 50 °C for three hours. Their genomic DNAs were then pooled in equal amounts for whole-exome sequencing at 200x average coverage (Novogene). As a control, genomic DNAs isolated from the corresponding cell populations before HQ461 selection were also pooled for whole-exome sequencing.
GEO Accession
GSM4649357