2.3 Bioinformatic and statistical analyses
DNA sequences were translated in-silico by the Translation Tool provided in the ExPASy portal to check for any pseudogenes and avoid sequencing errors. The composition of nucleotides was calculated on the CLC Workbench v7.7.1 and Mega X. The alignment of the amplicons was carried out in MUSCLE with the default specifications. Kimura two parameter (K2P) model was employed to calculate pairwise genetic distance at the level of species, genus and family (Kimura, 1980). Using this distance matrix, a Neighbor-Joining (NJ) algorithm was implemented to generate the phylogenetic tree topology in Mega X with a bootstrapping value of 1000 replications (Saitou and Nei, 1987, Felsenstein, 1985). Basic statistical analyses were done in Excel 2013 and the hypotheses testing (e.g.: t -test, F -test, etc.) was done in RStudio-1.2.5001.