Genetic techniques
Oligonucleotides used in this study were synthesized by Integrated DNA  Technologies (IDT; Coralville,  IA) and are listed in Table S2.Genomic DNAs (gDNAs) were sequencing using Illumina whole genome sequencing (SeqCenter; Pittsburgh, PA) and results were analyzed using BRESEQ software (Barrick et al., 2009). Plasmids used in this study are listed in Table S3 and were constructed using Gibson assembly as described previously (Gibson et al., 2009) and verified by Sanger sequencing (Azenta Life Sciences) or whole plasmid sequencing (Plasmidsaurus; Eugene, OR). E. coli NEB 5α (NEB) was used as a host for plasmid constructions, which were then transferred intoS. aureus strain RN4220 by electroporation as previously described (Schenk & Laddaga, 1992). Plasmids and marked  mutations  were  moved  between  S.  aureus strains via Φ85-mediated transduction (Novick, 1991).
Construction of the lpdA::kan+ strain:To construct the lpdA::kan+ strain (SRB3000), we moved the transposon insertion containing the KmRmarker from the S. aureus SH1000 strain VKS102 (Singh et al., 2008) into the S. aureus USA300 LAC SRB2421 (lpdA ::φNΣ) via Φ85-mediated transduction (Novick, 1991). Recombinants were selected on TSA Km medium and scored for Em susceptibility. We then confirmed the allele by PCR.