Genome sequencing and annotation Genome project history This organism was selected for sequencing on the basis of its phylogenetic position [25], and is part of the Genomic Encyclopedia of Bacteria and Archaea project [26]. The genome project is deposited in the Genome On Line Database [15] and the complete genome sequence full report is deposited in GenBank. Sequencing, finishing and annotation were performed by the DOE Joint Genome Institute (JGI). A summary of the project information is shown in Table 2. Table 2 Genome sequencing project information Growth conditions and DNA isolation H. hydrossis OT, DSM 1100, was grown in DSMZ medium 134 (Haliscomenobacter Medium) [27] at 26��C. DNA was isolated from 0.

5-1 g of cell paste using MasterPure Gram-positive DNA purification kit (Epicentre MGP04100) following the standard protocol as recommended by the manufacturer, with modification st/DL for cell lysis as described in Wu et al. [26]. DNA is available through the DNA Bank Network [28]. Genome sequencing and assembly The genome was sequenced using a combination of Illumina and 454 sequencing platforms. All general aspects of library construction and sequencing can be found at the JGI website [29]. Pyrosequencing reads were assembled using the Newbler assembler (Roche). The initial Newbler assembly consisting of 153 contigs in three scaffolds was converted into a phrap [30] assembly by making fake reads from the consensus, to collect the read pairs in the 454 paired end library. Illumina GAii sequencing data (1,273.3 Mb) was assembled with Velvet [31] and the consensus sequences were shredded into 1.

5 kb overlapped fake reads and assembled together with the 454 data. The 454 draft assembly was based on 369.3 Mb 454 draft data and all of the 454 paired end data. Newbler parameters are -consed -a 50 -l 350 -g -m -ml 20. The Phred/Phrap/Consed software package [30] was used for sequence assembly and quality assessment in the subsequent finishing process. After the shotgun stage, reads were assembled with parallel phrap (High Performance Software, LLC). Possible mis-assemblies were corrected with gapResolution [29], Dupfinisher [32], or sequencing cloned bridging PCR fragments with subcloning. Gaps between contigs were closed by editing in Consed, by PCR and by Bubble PCR primer walks (J.-F. Chang, unpublished).

A total of 589 additional reactions were necessary to close gaps and to raise the quality of the finished sequence. Illumina reads were also used to correct potential base errors and increase consensus quality using a software Polisher developed at JGI [33]. The error rate of the completed genome sequence is less than 1 in 100,000. Together, the combination of the Illumina Batimastat and 454 sequencing platforms provided 203.8 �� coverage of the genome. The final assembly contained 1,005,536 pyrosequence and 35,370,321 Illumina reads.