## 3 Results
### OsCM Overexpression Plant Construct and Phenotype Evaluation  
We conducted colony PCR of 10 selected colonies of transformed yeast and checked the transformation (Figure 3A). We the ascertained confirmation of cloning and transformation of the gene of interest in _Escherichia coli_ through restriction enzyme cutting. Colony PCR was used to select five colonies whose transformation was confirmed, and PCR was performed using _OsCM_ primer (Figure 3B). As a result of PCR, transformation was confirmed in all five colonies. Western blot confirmation of _OsCM_ expression in yeast. Similar amounts of protein (20 ug) isolated from transformed yeast strain and control strain after 60 h. Following 12% SDS-PAGE, the gels were stained with Coomassieie blue and  
Figure 2: Extraction of substances related to Cochlioquinone-9 (cq-9). (**A**) Molecular weight of cq-9. Mass: 381.38. (**B**) Chemical structure of cq-9; (3K)-9-[(1S,2R,3S)-2-acetyloxy-1,3-dimethylpentyl] 1,2,3,4a,5,6,6a,12,12a,12b-decahydro-12-hydroxy-3-(1-hydroxy-1-methylethyl)-6a,12b-dimethylpyrano[3,2-a]xanthene-8,11-dione, C\({}_{30}\)H\({}_{44}\)O\({}_{8}\), molecular weight = 532.7 g/mol. treated with an antibody for the recombinant protein (Figure 3C). The 13 kDa band was thickened in the yeast transformed by _OsCM_. The recombinant protein of the transformed strain is significantly expressed as compared with the wild type (Figure 3D). Furthermore, analysis of major secondary metabolites. Alanine, tyrosine, phenylalanine and alanine biochemical analysis. The wild type is a control, and the mutant is transformed with the _OsCM_. Alanine, tyrosine, phenylalanine and \(\beta\)-alanine were all highly expressed in _OsCM_-transformed yeast. In particular, we analyzed the OsCM functional expression in yeast and were reported the highest quantity of Phe (249 \(\mathrm{\SIUnitSymbolMicro mL}\)) and Tyr (108 \(\mathrm{\SIUnitSymbolMicro g}\)/mL) (Figure 3E). Supplementary data (Supplementary Figure S1) online describes the role of CM in shikimate pathway and correlation of shikimate pathway with cq-9. In this pathway, the key step is the conversion of chorismate into prephenate, which is the main precursor of aromatic amino acids. Chorismate mutase enzyme is involved in the conversion of chorismate into prephenate. Phenylalanine is the precursor of the phenylpropanol pathway, which is the intermediate of a high diversity of metabolites, including flavonoids and anthocyanins. The shikimate pathway produces salicylic acid (SA) in two ways; the iso-chorismate synthase enzyme converts anthraquinone into SA in the initial steps, while, in the last steps, phenylalanine undergoes a series of reactions and produces SA. SA is the ultimate source of catechol goes under various reaction, which is a key unit of cq-9. When WBPH was inoculated with OX-_OsCM_ and Cheongcheong, various phenotypes were analyzed. In OX-_OsCM_, lesions rarely form and appear very small in size, but in Cheongcheong, many lesions form and persist for a long period of time (Figure 4A-C). Especially, Cheongcheong had a long lesion length, but OX-_OsCM_ showed little lesion (Figure 4D-F). Moreover, Cheongcheong formed many lesions and died as the leaves  
Figure 3: Chorismate mutase (CM) molecular analysis. (**A**) Colony PCR of 10 selected colonies of transformed yeast on the 0.8% agarose gel. And target gene size is 378 bp. Lane; numbers 1 to 10 represent the selected colony number. M; \(\lambda/H\)_ind_III DNA ladder. (**B**) Confirmation of cloning and transformation of the gene of interest in _Escherichia coli_ through restriction enzyme cutting. The upper band represents the vector, whereas the lower band shows the target gene size (378 bp). (**C**) Western blot confirmation of CM expression in yeast. Similar amounts of protein (20 \(\mathrm{\SIUnitSymbolMicro g}\)) isolated from transformed yeast strain and control strain after 60 h. Following 12% SDS-PAGE, the gels were stained with Coomassieie blue and treated with an antibody for the recombinant protein.