ViralORFeome: an integrated database to generate a versatile collection of viral ORFs

Pellet, J.; Tafforeau, L.; Lucas-Hourani, M.; Navratil, V.; Meyniel, L.; Achaz, G.; Guironnet-Paquet, A.; Aublin-Gex, A.; Caignard, G.; Cassonnet, P.; Chaboud, A.; Chantier, T.; Deloire, A.; Demeret, C.; Le Breton, M.; Neveu, G.; Jacotot, L.; Vaglio, P.; Delmotte, S.; Gautier, C.; Combet, C.; Deleage, G.; Favre, M.; Tangy, F.; Jacob, Y.; Andre, P.; Lotteau, V.; Rabourdin-Combe, C.; Vidalain, P. O.

ViralORFeome database can be accessed at: http://www .viralorfeome.com. When selecting 'Virus' option in the menu bar, ViralORFeome web interface allows users to navigate throughout viral sequences retrieved from GenBank by entering criteria such as virus species, taxon IDs or accession numbers (Supplementary Figure S2 ). ViralORFeome can also execute a BLASTn or BLASTp search among available viral sequences (20) . Virus genomes and coding sequences of interest are visualized with an integrated genome browser adapted from GBrowse that provides a synthetic view of genomic sequence features (21) . When users have identified a viral ORF sequence that they want to clone, clicking on this sequence automatically generates cloning primers and virtual clones in ViralORFeome database ( Figure 2 ). ORF-specific Gateway Õ primers are designed using the Oligonucleotide Selection Program (OSP) software adapted for recombinational cloning (22) . By default, primers are designed to clone full-length ORFs from ATG to STOP codon according to GenBank annotations. However, users can specify 5 0 -and 3 0 -coordinates to clone ORF fragments corresponding to specific domains or even upload manually designed primers. Once primers have been validated, virtual clones are generated and used as entry points in ViralORFeome to store and access all information relative to the viral ORF constructs (e.g. primers, sequencing traces, potential mutations, and comments). Importantly, when users want to clone some variant of an ORF that is not defined in GenBank, and therefore is not available in ViralORFeome, an interface allows for the creation of a 'variant' clone that is anchored to the most similar annotated sequence found by BLAST in the database (Supplementary Figure S3 ). This ensures a maximum of flexibility when viral sequences are known and characterized but not readily available in GenBank, a frequent situation when working with primary isolates. This also facilitates the management of mutants generated on purpose to study the role of specific residues in a protein's function.