Computational protocol: Full Genome Sequencing and Genetic Characterization of Eubenangee Viruses Identify Pata Virus as a Distinct Species within the Genus Orbivirus

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Protocol publication

[…] ‘Raw’ ABI sequence data, was assembled into ‘contigs’ using the SeqManII sequence analysis package (DNAstar version 5). The ORFs of EUBV and TILV genome segments were identified using EditSeq and NCBI ORF finder, and translated to aa sequences for further analysis. Putative functions were assigned to individual proteins by comparisons to sequences in GenBank and identification of homologous BTV proteins, using BlastX (http://blast.ncbi.nlm.nih.gov/Blast.cgi?CMD=Web&PAGE_TYPE=BlastHome). Multiple alignments of consensus sequences were performed using Clustal X (Version 2.0) , Clustal Omega (http://www.ebi.ac.uk/Tools/msa/clustalo/) and MAFFT (http://mafft.cbrc.jp/alignment/server/) to ensure proper alignment. Pairwise distance (aa and nt) calculations and phylogenetic trees constructions were done using MEGA 5 software with the p-distance parameter and neighbor-joining method . The hydrophobicity profile of different NS4 proteins was analysed using the Kyte and Doolittle hydrophobicity plot with a window size of 11 amino acids (aa) . Sequence relatedness to proteins and domains were assessed by comparing with the pfam database (Available at http://pfam.sanger.ac.uk/search). The presence of ‘coiled-coils’ was analysed using the program ‘COILS’ (available at http://www.ch.embnet.org/software/COILS_form.html). Nuclear localisation signals were predicted using cNLS Mapper (available at http://nls-mapper.iab.keio.ac.jp/cgi-bin/NLS_Mapper_form.cgi). […]

Pipeline specifications

Software tools Open Reading Frame Finder, BLASTX, Clustal W, Clustal Omega, MAFFT, MEGA, cNLS Mapper
Databases Pfam
Applications Phylogenetics, Protein sequence analysis
Organisms Human poliovirus 1 Mahoney, Viruses
Chemicals Amino Acids