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Additional file 1: Fig. S1. O. pumila plant materials were used for metabolite profiling. Fig. S2. Analysis of gene expression patterns and phylogenic analysis of STRs, SLSs, LAMTs enzymes in CPT biosynthesis. Fig. S3. Protein sequence alignments mentioned in this article. Fig. S4. OpLAMT assay with secologanic acid and OpSTR competion expriments. Fig. S5. The standard curve of strictosidine and strictosidinic acid. Fig. S6. The SDS-PAGE gel of purified recombinant proteins used in the chemo-enzymatic synthesis of deuterium-labeled metabolites and biochemical assay. Fig. S7. Scheme of labeled substrates synthesis. Fig. S8. Chemoenzymatic synthesis of labeled substrates. Fig. S9. Detection of d4-strictosidinic acid and d4-strictosidine in the feeding experiments in O. pumila. Fig. S10. Metabolites detection of N. nimmoniana by LC-MS. Fig. S11. Phylogenetic relationship of DNA topoisomerase I in CPT-producing and non-producing species. Table S1. Relevant Compounds Detected in O. pumila plant and hairy root. Table S2. Identification of candidate CPT biosynthetic pathway genes in O. pumila as revealed by sequence identity with characterized genes from the pre-strictosidine biosynthetic pathways in Catharanthus roseus. Table S3. Identities and similarities among STRs from C. acuminata, N. nimmoniana, and O. pumila used in this work. Table S4. Kinetic parameters of OpSTR towards secologanin and secologanic acid. Table S5. Primers list used in this study.

Additional file 2:. The original, uncropped SDS-PAGE of Fig. S6.

qRT-PCR expression analyses of HsLDC, HsCAO, HsPKS, and CYP450s. (PDF 36Â kb)

Summary of H. serrata RNA seq by two Next-Generation Sequencing Systems (PDF 44Â kb)

Unrooted neighbor-joining phylogenetic tree of selected CYP450s (Bootstrap values (1000 replicates)). (PDF 45Â kb)

The ORF or amino acid sequences of three types (LDC, CAO, PKS) of enzymes identified from H. serrata and found in the transcriptome. LDC (HsLDC-X1, HsLDC-X2, HsLDC-X3, HsLDC-X4, HsLDC-X5, HsLDC-X6, AB915697.1, AB915696.1, Unigene94617, Unigene94988); CAO (HsCAO, CL4248.1, CL4248.2, CL4248.3); PKS (HsPKS1 (ABI94386.1), HsPKS2, HsPKS3, Unigene393, Unigene394, CL2724.2). (PDF 63Â kb)

The amino acid sequences of 12 homologues identified as SLS. (PDF 35Â kb)

Selected genes for LDC’s, CAO’s, and PKS’s phylogenetic analysis. (PDF 66 kb)

The amino acid sequences of 11 homologues identified as BBE. (PDF 33Â kb)

LDC’s, CAO’s, and PKS’s multiple-tissues specific expression patterns analysis by heat map. (PDF 63 kb)