Computational protocol: Perspectives on the Evolution of Porcine Parvovirus

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Protocol publication

[…] For the phylogenic analysis, the complete sequences were downloaded from GenBank and aligned using the ClustalW program in the BioEdit software version 7.0.9 []. Phylogenic trees were inferred by the maximum-clade credibility method (nucleotide sequences) implemented in Beast version 1.8.2. The branches of the maximum clade credibility tree were colored according to the most probable location state of their descendent nodes. To estimate the substitution rates per site per year and the time in the NS1, VP1 and VP2 genes of PPV, we aligned 71 complete NS1, 65 complete VP1, and 75 VP2 sequences from GenBank and aligned by using the ClustalW program in the software BioEdit software version 7.0.9 []. Root-to-tip analysis was conducted using TemPest version 1.5 to assess whether there was sufficient temporal signal to proceed with the phylogenic molecular clock analysis. Sequences that were not suitable for analysis were excluded in the molecular clock analysis []. Rates of nucleotide substitutions per site per year and time to most recent common ancestor (TMRCA) were estimated using the Bayesian framework [], which was applied to reconstruct the spatial-temporal diffusion history of PPV. In brief, the spatial diffusion of the time-scaled genealogy is modeled as a standard continuous-time Markov chain (CTMC) process over discrete sampling locations. A Bayesian stochastic search variable selection (BSSVS) approach, which allows the exchange rates in the CTMC to be zero with some prior probability, was used to find a parsimonious set of rates explaining the diffusions in the phylogeny. The analysis was performed using Beast package v1.8.2 under the following assumptions (i) a codon based SRD06 nucleotide substitution model, (ii) a constant population size for the coalescent prior, and (iii) the molecular clock model of uncorrelated lognormal distribution. The analysis was run for 100 million chains, sampling every 10,000 generations. The phylogenic trees were summarized with TreeAnnotator and were depicted using FigTree []. Groupings with posterior probabilities over 0.90 were considered to be clusters and those with posterior probabilities less than 0.90 were considered to be clades. This process was also performed using 75 sequences of the complete VP2 gene and 71 sequences of the complete NS1 gene (). [...] For the detection of potential recombination events, we aligned the complete NS1, VP1, and VP2 sequences of 42 PPV strains and used a recombination detection program (RDP version 4.460). X-over automated RDP analysis was used to identify recombination points within the PPV genome. For estimates of amino acid mutations, we compared the similarity of 75 VP2 sequences with the T142_South Korea strain using the DNAstar (Lasergene, Madison, WI, USA) program and aligned each of sequences using the ClustalW program in BioEdit version 7.0.9 []. […]

Pipeline specifications

Software tools Clustal W, BioEdit, BEAST, TempEst, FigTree, RDP4, DNASTAR Lasergene
Applications Phylogenetics, Sanger sequencing
Organisms Porcine parvovirus, Sus scrofa, Plum pox virus
Diseases Parvoviridae Infections
Chemicals Amino Acids