Compositions and methods for making (r)-reticuline and precursors thereof

Abstract

Methods that may be used for the manufacture of the chemical compound (R)-Reticuline and synthesis precursors thereof. Compositions useful for the synthesis (R)-Reticuline and synthesis precursors are also provided.

1 - 84 . (canceled) 
     
     
         85 . A recombinant nucleic acid comprising a nucleic acid sequence at least 70% identical to one or more of the nucleic acid sequences selected from the group consisting of:
 (i) SEQ ID NO: 116-218, SEQ ID NO: 324, and SEQ ID NO: 337;   (ii) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 219-321, SEQ ID NO: 325, and SEQ ID NO: 338;   (iii) SEQ ID NO: 1-58, SEQ ID NO: 326, SEQ ID NO: 328, and SEQ ID NO: 339;   (iv) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 59-115, SEQ ID NO: 327; SEQ ID NO: 329, SEQ ID NO: 330, and SEQ ID NO: 340;   (v) SEQ ID NO: 322; and   (vi) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 323.   
     
     
         86 . The recombinant nucleic acid according to  claim 85 , wherein the nucleic acid is a complementary DNA (cDNA). 
     
     
         87 . A chimeric nucleic acid sequence comprising:
 (a) a first nucleic acid sequence encoding a CYP450 polypeptide; and   (b) a second nucleic acid sequence encoding an AKR polypeptide; and   (c) one or more nucleic acid sequences capable of controlling expression in a host cell.   
     
     
         88 . The chimeric nucleic acid sequence according to  claim 87 , wherein the first nucleic acid sequence comprises a nucleic acid sequence at least 70% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO: 116-218, SEQ ID NO: 324, and SEQ ID NO: 337. 
     
     
         89 . The chimeric nucleic acid sequence according to  claim 87 , wherein the second nucleic acid sequence comprises a nucleic acid sequence at least 70% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO: 1-58, SEQ ID NO: 326, SEQ ID NO: 328, and SEQ ID: NO: 339. 
     
     
         90 . The chimeric nucleic acid sequence according to  claim 87 , wherein (i) the first nucleic acid sequence comprises a nucleic acid sequence at least 70% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO: 116-218, SEQ ID NO: 324, and SEQ ID NO: 337, and (ii) the second nucleic acid sequence comprises a nucleic acid sequence at least 70% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO: 1-58, SEQ ID NO: 326, SEQ ID NO: 328, and SEQ ID: NO: 339. 
     
     
         91 . The chimeric nucleic acid sequence according to  claim 87 , wherein the nucleic acid sequence capable of controlling expression comprises a promoter, a transcriptional terminator, a transcriptional enhancer, or combinations thereof. 
     
     
         92 . A recombinant expression vector comprising as operably linked components:
 (a) a nucleic acid sequence capable of controlling expression in a host cell; and   (b) a nucleic acid sequence encoding CYP450 and AKR;   wherein the expression vector is suitable for expression in a host cell.   
     
     
         93 . The recombinant expression vector according to  claim 92 , wherein the nucleic acid sequence encoding CYP450 comprises a nucleic acid sequence at least 70% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO: 116-218, SEQ ID NO: 324, and SEQ ID NO: 337. 
     
     
         94 . The recombinant expression according to  claim 92 , wherein the nucleic acid sequence encoding AKR comprises a nucleic acid sequence at least 70% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO: 1-58, SEQ ID NO: 326, SEQ ID NO: 328, and SEQ ID NO: 339. 
     
     
         95 . The recombinant expression vector of  claim 92 , wherein the nucleic acid sequence encoding CYP450 and AKR comprises at least 70% identical to the nucleic acid sequence of SEQ ID NO: 322. 
     
     
         96 . The recombinant expression vector of  claim 92 , wherein the nucleic acid sequence capable of controlling expression in a host cell comprises a promoter, a transcriptional terminator, a transcriptional enhancer, or combinations thereof. 
     
     
         97 . The recombinant expression vector of  claim 92 , further comprising one or more nucleic acid sequences encoding marker genes, one or more origins of replication, genetic elements required for the integration of the vector or a portion thereof into the host cell's genome, or combinations thereof. 
     
     
         98 . A host cell comprising a nucleic acid sequence according to claim  1 . 
     
     
         99 . The host cell of  claim 98 , wherein the host cell is a non-plant cell. 
     
     
         100 . The host cell of  claim 99 , wherein the host cell is  Escherichia coli.    
     
     
         101 . The host cell of  claim 99 , wherein the host cell is a yeast cell.