Transposon Insertion Site Identification

MAP mutants were lysed in the deionized water with mechanical disruption method using 0.2 mL of 0.1-mm-diameter glass beads. Samples were cleared by centrifugation at 10,000× g for 5 min and DNA was purified using a DNA Clean and Concentrate kit (Zymo Research, Irvine, CA, USA) following the manufacturer’s protocol. The transposon insertion site was identified using the previously reported ligation-mediated polymerase chain reaction (LM-PCR) technique that is comprehensively described in [24 (link)]. The final PCR products were separated on the agarose gel, and purified DNA fragments sequenced at the Center for Genome Research and Biocomputing, Oregon State University.