Shotgun sequencing of induced phage lysate from Escherichia coli strain HS: mitomycin C induction, mixed virion DNA including transducing phage
Virion DNA extracted from induced prophage particles. Escherichia coli strain HS was treated with mitomycin C (0.5 ug/mL) for h. The lysate was treated with DNase I and RNase A (2 ul each) for 30min at 37C to degrade unencapsidated nucleic acids. Isolation of virion DNA was performed via PEG Method using PEG (polyethylene glycol) and NaCl. Libraries for wholegenome sequencing were prepared from 500 ng of phage DNA using the MGI Easy PCRFree Library Prep Set according to the manufacturer's instructions. Enzymatic fragmentation was performed as recommended, followed by size selection of 400450 bp fragments using the provided DNA Clean Beads. Library concentration was measured using a Qubit Flex fluorometer with the dsDNA HS Assay Kit. Library quality was assessed with a 4200 TapeStation System using the High Sensitivity D1000 ScreenTape Assay. Libraries were circularized, pooled and sequenced on a DNBSEQG400 platform in 2  150 bp pairedend mode.