Genome Sequencing of Listeria monocytogenes

Both L. monocytogenes strains were cultivated under aerobic conditions at 37°C in brain heart infusion broth (BHI, Merck; with 125 rpm shaking) and harvested by centrifugation. The resulting pellet was used for DNA isolation using the QIAGEN genomic-tip columns and buffers (QIAGEN), in accordance with the recommendations of the manufacturer. Genome sequencing was performed using an Illumina GAII genome analyzer at the University of Veterinary Medicine Vienna, Austria. Sequencing was performed using paired-end sequencing technology and 101 bp read-length using Illumina standard protocols. Ten million reads were used for a de novo assembly using SeqManNGen (DNASTAR). For strain QOC1, the assembly of sequenced data resulted in generation of 17 contigs (>500 bp) for which there was an average coverage of 310×, while for strain QOC2, assembly of the reads resulted in 13 contigs (>500 bp), for which the average coverage was 330×. For ordering, the contigs were aligned to the L. monocytogenes EGDe genome using the “move contigs” option in MAUVE [22] (link). Some of the remaining gaps could be closed by PCR and Sanger sequencing - resulting in seven and four contigs in the final assembly for strain QOC1 and QOC2, respectively.