Dendrogram analysis and protein sequence alignment

A nucleotide sequence alignment of the poplar nitrilase genes and nitrilase genes from other plants was constructed with the Muscle (codon) algorithm (gap open, − 2.9; gap extend, 0; hydrophobicity multiplier, 1.5; clustering method, upgmb) implemented in MEGA6 [39]. The tree was reconstructed with MEGA6 using the Maximum Likelihood method (model/method, Kimura-2-parameter model; substitutions type, nucleotide; rates among sites, uniform rates; gaps/missing data treatment, partial deletion; site coverage cutoff, 80%) and a bootstrap resampling analysis with 1000 replicates was performed to evaluate the tree topology. An amino acid alignment of the poplar nitrilases was constructed and visualized with BioEdit (http://www.mbio.ncsu.edu/bioedit/bioedit.html) and the ClustalW algorithm.