Genome Sequencing of 34 Synechococcus Strains

Thirty-four Synechococcus strains were chosen for genome sequencing based on their phylogenetic position, pigment content and isolation sites (Figure 1 and Supplementary Table S1). All but the three KORDI strains were retrieved from the Roscoff Culture Collection (RCC²) and transferred three times on 0.3% SeaPlaque Agarose (Lonza, Switzerland) to clone them and reduce contamination by heterotrophic bacteria. A first set of 25 Synechococcus genomes (including WH8103) were generated at the Genoscope (CEA, Paris-Saclay, France) by shotgun sequencing of two libraries: a short-insert forward-reverse pair-end (PE) library (50–150 bp) and a long-insert reverse-forward mate-pair library (4–10 kb), both sequenced by Illumina^TM technology. Additionally, seven other genomes were sequenced at the Center for Genomic Research (University of Liverpool, United Kingdom) by shotgun sequencing of 250 bp reads. Single or PE reads were first assembled into contigs using the CLC Assembly Cell© 4.10 (CLC Bio, Aarhus, Denmark). Synechococcus contigs were identified based on their different coverage compared to heterotrophic bacteria, scaffolded using WiseScaffolder and 28 out of 31 genomes were closed by manual finishing as described in Farrant et al. (2015) (link). Three genomes (BIOS-E4-1, BOUM118, and RS9915), had only one to three gaps in highly repeated genomic regions. The base numbering of the circularized genomes was set at 174 bp before the dnaN start, corresponding approximately to the origin of replication. Automatic structural and functional annotation of the genomes was then realized using the Institute of Genome Science (IGS) Annotation Engine³ (Galens et al., 2011 (link)). As concerns KORDI-49, KORDI-52 and KORDI-100 strains, genomes were sequenced from axenic cultures using a 454 GS-FLX Titanium sequencing system (Roche) at Macrogen (Seoul, South Korea). The obtained reads were assembled using the Newbler assembler (version 2.3, Roche). To fill contig gaps, additional PCR and primer walking was conducted. Sequences of all new Synechococcus genomes were deposited in GenBank under accession numbers CP047931-CP047961 (BioProject PRJNA596899), except Synechococcus sp. WH8103 that was previously deposited to illustrate the performance of the pipeline used to assemble, scaffold and manually finish these genomes as well as the three KORDI genomes that have been deposited in Genbank in August 2014 (see accession numbers in Supplementary Table S1).