Candidates for AmISY were identified based on sequence homology to ISY from C. roseus (CrISY) in a genome sequence of the JI7 inbred line of A. majus (http://snapdragon.genomics.org.cn/).4 The protein sequence of CrISY was used in a BLAST search against proteins predicted from the genome sequence to yield four hits with amino acid sequence identities between 39% and 66% (Fig. 2a). Candidate Am18679 showed the highest amino acid similarity to CrISY, with 66% identity and 79% similarity. For overexpression in Escherichia coli, all four genes were cloned from cDNA of A. majus flower and leaf tissue and successfully purified via nickel affinity chromatography. Enzyme reactions containing 8-oxogeranial and NADPH as substrates were analyzed by GC-MS. Only protein derived from candidate Am18679 (Fig. 2b), the candidate most similar to CrISY, yielded sizeable quantities of cyclized iridoid product (supplemental Fig. S1). Therefore, candidate Am18679 was named AmISY. Only trace amounts of substrate were consumed, and negligible products were detectable with the more distantly related candidates.