The initial draft assembly contained 199 contigs in 5 scaffolds. The 454 paired end data was assembled with Newbler, version 2.3. The Newbler consensus sequences were computationally shredded into 2 kb overlapping fake reads (shreds). Illumina sequencing selleck catalog data were assembled with VELVET, version 1.0.13 , and the consensus sequence were computationally shredded into 1.5 kb overlapping fake reads (shreds). We integrated the 454 Newbler consensus shreds, the Illumina VELVET consensus shreds and the read pairs in the 454 paired end library using parallel phrap, version SPS – 4.24 (High Performance Software, LLC). The software Consed [42-44] was used in the following finishing process. Illumina data was used to correct potential base errors and increase consensus quality using the software Polisher developed at JGI (Alla Lapidus, unpublished).
Possible mis-assemblies were corrected using gapResolution (Cliff Han, unpublished), Dupfinisher (Han, 2006), or sequencing cloned bridging PCR fragments with subcloning. Gaps between contigs were closed by editing in Consed, by PCR and by Bubble PCR (J-F Cheng, unpublished) primer walks. A total of 275 additional reactions were necessary to close gaps and to raise the quality of the finished sequence. The estimated genome size is 7.6 Mb and the final assembly is based on 65.3 Mb of 454 draft data which provides an average of 8.6�� coverage of the genome and 4,864.7 Mb of Illumina draft data which provides an average 640.1�� coverage of the genome.
Genome annotation Genes were identified using Prodigal  as part of the DOE-JGI Annotation pipeline , followed by a round of manual curation using the JGI GenePRIMP pipeline . The predicted CDSs were translated and used to search the National Center for Biotechnology Information (NCBI) non-redundant database, UniProt, TIGRFam, Pfam, PRIAM, KEGG, COG, and InterPro databases. These data sources were combined to assert a product description for each predicted protein. Non-coding genes and miscellaneous features were predicted using tRNAscan-SE , RNAMMer , Rfam , TMHMM , and SignalP . Additional gene prediction analyses and functional annotation were performed within the Integrated Microbial Genomes (IMG-ER) platform [37,53]. Genome properties The genome is 8,618,824 nucleotides with 60.74% GC content (Table 3) and comprised of 32 contigs in 6 scaffolds (Figure 3).
From a total of 8,576 genes, 8,493 were protein encoding and 83 RNA only encoding genes. The majority of genes (77.85%) were assigned a putative function whilst the remaining genes were annotated as hypothetical. AV-951 The distribution of genes into COGs functional categories is presented in Table 4. Table 3 Genome sequencing project information for Rhizobium leguminosarum bv. trifolii strain SRDI943. Figure 3 Graphical linear map of the genome of Rhizobium leguminosarum bv. trifolii strain TA1.