Fig. 4

Identification of key genes of downstream of steroidal saponins biosynthetic pathway (DSSP); A, the phylogenetic tree was constructed using CYP450s of asparagus and the known CYP90s family genes involved in steroids C22 hydroxylation based on protein sequence similarity by Neighbor-Joining Algorithm (NJ), and the gene name listed as gene symbol, organism name and NCBI access No., respectively, while the asparagus genes showed expression level with the heatmap in GR, GS, GF, PR, PS, and PF with 3 replicates, respectively; B, the multiple sequence alignment of three CYP90s genes in asparagus with functionally characterized CYP90s genes, highlighting the conserved amino acid residues in red; C, the phylogenetic tree of CYP450s from asparagus and known CYP72s and CYP94s family involved in steroids C26 hydroxylation based on protein sequence similarity by NJ, the gene name listed as above; D, the phylogenetic tree was constructed using genes annotated as glucosidase in asparagus and functionally characterized furostanol glycoside 26-O-beta-glucosidase (F26G) genes by NJ, with gene name followed with their protein conserved motifs predicted using MEME. E, the phylogenetic tree was constructed using glycosyltransferases (GTs) genes in asparagus and functionally characterized GTs genes by NJ and the motifs were shown as in D