title: Tn-seq of MAB during infection
design: Samples were prepared for transposon sequencing following previously published protocol (see Tn-Seq Illumina Library Preparation from Initial PA14 Mutant Library in supplemental material for PMID: 25775563) with the following modifications: 1) MycoMar specific primers were used see Table S17, 2) between steps DNA was purified using the GeneJet PCR purification kit (Thermo Scientific, Waltham, MA); 3) 2-4 PCR-1 reactions were performed, each with 0.5 to 1 g DNA and these reactions were pooled in the following purification step; and 4) a mixture of 4 mariner-2 primers were combined with 3N, 5N, 7N, and 9N random bases to increase the sequence diversity of our libraries, 5) gel based size exclusion after PCR-2 using following the NEB QC Check and Size Selection using 6% PolyAcrylamide Gel - NEBNext Multiplex Small RNA Sample Prep Set for Illumina protocol. DNA was cut out between 160bp and 307bp93.  Libraries were pooled and sequenced at the Georgia Tech Molecular Evolution Core using a NextSeq 500 sequencer (Illumina, San Diego, CA) with 1  75 v2 chemistry.