Whole-cell lysis protocol

OA Ozan Alkan
BS Birgit Schoeberl
MS Millie Shah
AK Alexander Koshkaryev
TH Tim Heinemann
DD Daryl C. Drummond
MY Michael B. Yaffe
AR Andreas Raue
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Cells were grown and treated in a 6-cm dish scale. To harvest cells, medium was removed and quickly replaced by ice-cold PBS. PBS was then replaced with 250 μl of 2% SDS lysis buffer. After 5 min of incubation, the lysed cells were scraped off and loaded onto a cell lysate homogenizer microcentrifuge spin-column (catalog no. 79656, QIAshredder, Qiagen). The filtrate was then loaded onto a 0.2-m centrifugal filter column [catalog no. ODM02C34, Nanosep MF (0.2 m), Pall]. The lysates were then stored at −80°C. SDS-containing lysis buffer (2%) was modified after (44).

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