Systems, methods, and compositions for high-level production of water-soluble cannabinoids in a plant trichome
The inventive technology relates to systems and methods for enhanced in vivo production, accumulation and modification of cannabinoids. In one embodiment, the invention may include systems and methods for enhanced in vivo biosynthesis of chemically-modified water-soluble cannabinoids in a whole plant, or a cell suspension culture system.
1. A method for producing water-soluble cannabinoids in a trichome comprising:
1) introducing into a Cannabis plant a first nucleotide sequence, operably linked to a promoter, encoding a fusion peptide having a first domain and a second domain, wherein said first domain comprises a N-terminal cannabinoid synthase trichome targeting domain, and said second domain comprises a glycosyltransferase enzyme, and wherein said glycosyltransferase enzyme is not endogenous to Cannabis , and is selected from the group consisting of: nucleotide sequence according to SEQ ID NO's. 7, 24, 27, 29, 31, 33, 35, 37, and 39;
2) introducing into the Cannabis plant a second nucleotide sequence, operably linked to a promoter, encoding a heterologous UDP-galactose/UDP-glucose transporter having a plasma membrane targeting sequence configured to facilitate the transport of UDP-glucose to the trichome of the Cannabis plant where it is used as a substrate for said glycosyltransferase enzyme, and wherein said plasma membrane targeting sequence replaces or disrupts the tonoplast targeting sequence of the UDP-galactose/UDP-glucose transporter; and
wherein said N-terminal cannabinoid synthase trichome targeting domain is selected from the group consisting of: an N-terminal cannabidiolic acid (CBDA) synthase trichome targeting sequence; an N-terminal cannabigerolic acid (CBGA) synthase trichome targeting domain; an N-terminal cannabichromenic acid (CBCA) synthase trichome targeting domain; and an N-terminal tetrahydrocannabinolic acid (THCA) synthase trichome targeting domain.
2. The method of claim 1 wherein the first domain of said fusion peptide is selected from the group consisting of: the amino acid sequence identified as SEQ ID NO. 40, and the amino acid sequence identified as SEQ ID NO. 41.
3. The method of claim 1 wherein said fusion peptide comprises nucleotide sequence SEQ ID NO. 19.
4. The method of claim 1 wherein said plasma membrane targeting sequence comprises the nucleotide sequence according to SEQ ID NO. 74.
5. The method of claim 1 wherein said heterologous UDP-galactose/UDP-glucose transporter having a plasma membrane targeting sequence comprises the nucleotide sequence according to SEQ ID NO. 21.