Towards the Description of the Genome Catalogue of Pseudomonas sp. Strain M1.

Pseudomonas sp. strain M1 is a soil isolate with remarkable biotechnological potential. The genome of Pseudomonas sp. M1 was sequenced using both 454 and Illumina technologies. A customized genome assembly pipeline was used to reconstruct its genome sequence to a single scaffold.

GENOME ANNOUNCEMENT

Pseudomonas sp. strain M1, isolated from the Rhine River (1), is able to utilize several toxic and/or recalcitrant compounds, such as myrcene (1, 2), citral, citronellol, phenol (3–5), chlorophenols, and benzene (5), as its sole carbon and energy sources. However, the molecular mechanisms of M1 strain that are associated with the utilization of those (and other) less common carbon sources are still poorly known. To set the proper background for exploring the biotransformation potential of Pseudomonas sp. M1, its genome was sequenced using both 454 FLX and Illumina Genome Analyzer IIx next-generation sequencing technologies. The 454 FLX sequencing technology yielded a 264,177 single-read data set with an average read length of 523 bp, whereas the Illumina technology was used to produce two 50-bp read-length data sets: (i) 5,303,579 pair-end reads with an estimated insert size of about 320 bp; and (ii) 5,478,608 mate-paired reads with an estimated insert size of about 5,200 bp. The removal of adapter sequences and quality trimming were performed in all data sets prior to de novo assembly. To reconstruct the genome of Pseudomonas sp. M1, a customized pipeline was set, based on preliminary comparative trials using different genome assemblers. First, 454 FLX single reads were assembled with Newbler v2.6 (6), generating 379 contigs (minimum contig size of 1,000 bp) with a total size of 6,860,386 bp. Second, the 454 FLX-generated contigs were used as a genome backbone to produce eight scaffolds using both Illumina libraries in SSPACE v2.0 (7). The scaffolds we obtained included over 200 gaps, which were significantly filled by combining local alignment with GapFiller (8) and GapCloser (9). To obtain a more contiguous and accurate genome sequence, a further sequential run of SSPACE (7), GapFiller (8) and GapCloser (9), and Anchor v0.3.1 (http://www.bcgsc.ca/platform/bioinfo/software/anchor) was done, resulting in a single scaffold representing the genome of Pseudomonas sp. M1. Further genome sequence accuracy was improved by manual curation (based on read alignment analysis) and Sanger sequencing to confirm sequence accuracy and to close different sequence gaps. Nonetheless, nine repeat-rich regions were not fully resolved. As a whole, the current draft of the Pseudomonas sp. M1 genome is composed of nine contigs organized in a single scaffold, with a total size of 6,958,606 bp (including 1,753 N's), with an estimated G+C content of 67.3%. This genome sequence was annotated using Prokka v1.5.2 (http://www.vicbioinformatics.com/software.prokka.shtml) and deposited at DDBJ/EMBL/GenBank. The annotated genome includes 6,053 coding sequences (CDSs), 12 rRNAs (four copies each of 5S, 16S, and 23S rRNA), 66 tRNAs, and 804 hypothetical proteins.

Further inspection of the genome of Pseudomonas sp. M1 revealed the presence of a significant number of biotechnologically interesting enzymes (e.g., 74 oxygenases/hydroxylases) whose functionality may be fine-tuned using systems and/or synthetic biology approaches.

Nucleotide sequence accession numbers.

This Whole Genome Shotgun project (Bioproject: PRJNA62721) has been deposited at DDBJ/EMBL/GenBank under the accession number ANIR00000000. The version described in this article is the first version, ANIR01000000.