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.