The second library (MAI1-BLS256) was obtained, using Xoo MAI1 as a ‘tester’ and Xoc BLS256 as a ‘driver’. SSH was performed, using the BD PCR-Select™ Bacterial Genome Subtraction Kit (BD Biosciences Clontech, Mountain View, CA). Briefly, genomic DNAs of the three bacterial strains were isolated

using the Wizard® Genomic DNA Purification Kit according to the manufacturer’s recommendations (Promega Corporation, Madison, WI). The genomic DNAs were separately digested with RsaI restriction endonuclease (New England this website Biolabs® Inc., Beverly, MA). The DNA subtracted from each library was directly inserted by TA cloning, using the pGEM®-T Easy Vector System (Promega Corporation), and transformed into chemically competent cells (TurboCells® Competent E. coli), as described by the manufacturer (Genlantis Inc., San Diego, CA). We randomly selected

2112 and 2304 individual colonies for the MAI1-PXO86 and MAI1-BLS256 SSH libraries, respectively. Plasmid DNA of subtracted library colonies was obtained from individual clones, using the alkaline lysis procedure according to R.E.A.L. Prep 96 protocols (Qiagen, S.A., Courtaboeuf, France). Insert sequences in the subtracted libraries were one-end sequenced with the T7 primer. We used the computational sequence analysis pipeline created by (Lopez et al., 2004) for cleaning raw sequences, contig construction, and sequence analysis, allowing automatic treatment of our data. This pipeline manages treatment Selleckchem Alectinib of the sequence from the raw sequence (chromatogram) to the creation

of a set of nonredundant the sequences. Bases were called, using the phred program (Ewing et al., 1998). End sequences were trimmed for low quality, and vector sequences were eliminated. Only sequences longer than 100 bp after this trimming process were included in the dataset. The stackpack™ software (Miller et al., 1999) was used to create a set of nonredundant sequences. In a first step, the stackpack™ program creates clusters of sequences having >96% identity over a window of 150 bases. In a second step, sequences from a cluster are assembled using the phrap program. The SSH Xoo MAI1 nonredundant set of sequences was deposited at GenBank’s GSS database (http://www.ncbi.nlm.nih.gov/dbGSS/) under accession numbers FI978060–FI978198. Sequences were searched against the NCBI database with blastn (http://blast.ncbi.nlm.nih.gov). blast searches were performed against the complete nonredundant database and the genomes of Xoo strains KACC10331, MAFF311018, and PXO99A; Xoc strain BLS256; Xanthomonas campestris pv. campestris (Xcc) strain ATCC 33913; Xanthomonas axonopodis pv. vesicatoria (Xav) strain 85-10; and X. axonopodis pv. citri (Xac) strain 306 using the default parameters.