title: Tn-seq from E. coli in a priB knockout strain with sulB103 allele. Replicate C
design: Genomic DNA was purified using a Wizard Genomic DNA Purification Kit (Promega) and quantified using the QuantiFluor ONE dsDNA System (Promega). Genomic DNA was sheared to ~200 bp fragments with sonication. The resulting gDNA fragments were prepared for sequencing using NEBNext Ultra II DNA Library Prep Kit for Illumina (New England Biolabs). Bead-based size selection was used to enrich for 200 bp fragments prior to a 21-cycle splinkerette PCR utilizing a custom Tn5-enriching forward primer (oAM055) and custom indexed reverse primers for multiplexing (oAMrev) [25]. To ensure the quality and length of amplified DNA, a final bead-based size selection was employed. DNA was then sequenced with a NextSeq platform (Illumina) at the University of Michigans Advanced Genomics Core using a custom read primer (oAM058) to read the last 10 nt of the transposon before entering chromosomal DNA (to ensure reads corresponded to Tn5-insertions). To maintain sufficient sequence diversity on the flow-cell, a PhiX DNA spike was also included in the run. A custom index read primer (oAM059) and standard Illumina primer (oAM112) were employed for sequencing.