LOCUS       sequence-344752-        8327 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Biopterin production pathway with J306 scRNA on pBBR1-GmR plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8327)
  AUTHORS   Kiattisewee C, Dong C, Fontana J, Sugianto W, Peralta-Yahya P, 
            Carothers JM, Zalatan JG
  TITLE     Portable bacterial CRISPR transcriptional activation enables 
            metabolic engineering in Pseudomonas putida.
  JOURNAL   Metab Eng. 2021 Apr 27. pii: S1096-7176(21)00063-X. doi: 
            10.1016/j.ymben.2021.04.002.
  PUBMED    33930546
REFERENCE   2  (bases 1 to 8327)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Metab Eng. 
            2021 Apr 27. pii: S1096-7176(21)00063-X. doi: 
            10.1016/j.ymben.2021.04.002."
COMMENT     Sequence Label: pPPC027.306
FEATURES             Location/Qualifiers
     source          1..8327
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(21..55)
                     /label=BBa_J23117
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.06"
     protein_bind    348..369
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(384..389,390..407,408..414)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    422..438
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     427..449
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     446..462
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     446..462
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     481..501
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        483..501
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     terminator      complement(546..592)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     745..762
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     CDS             complement(818..835)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(851..880)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     misc_RNA        complement(903..921)
                     /label=MS2 stem loop
                     /note="stem loop that binds the bacteriophage MS2 coat 
                     protein"
     promoter        complement(1005..1039)
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     primer_bind     complement(1053..1072)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1054..1072)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1082..1099)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1082..1098)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1091..1113)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             complement(1807..2469)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      2470..3241
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     CDS             complement(4523..5056)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(5245..5250,5251..5267,5268..5273))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     terminator      complement(5439..5466)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(5482..5553)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(6426..6443)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             6453..6464
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(6474..6508)
                     /label=BBa_J23117
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.06"
     terminator      complement(6688..6715)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(6731..6802)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(7249..7266)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             7276..7287
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(7297..7331)
                     /label=BBa_J23117
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.06"
     terminator      complement(7511..7538)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(7554..7625)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(8300..8317)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             join(8327,1..11)
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
ORIGIN
        1 ttctcctctt taatgaattc gctagcacaa tccctaggac tgagctagct gtcaatctat
       61 aatcgcaact tcaagacgac gttgtgtcca gaacgctccg taggagaagt gaggagacga
      121 gcgaacgcgt cgtacgagct ttatgcatct ttcacagctc gtagcgatga ttatgacatc
      181 gcagtgagca actgaataag cgctgcgtga atgatcgcaa atgctgatat cggaattgcc
      241 atgcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg
      301 cacgacaggt ttcccgactg gaaagcgggc agtgagcgca acgcaattaa tgtgagttag
      361 ctcactcatt aggcacccca ggctttacac tttatgcttc cggctcgtat gttgtgtgga
      421 attgtgagcg gataacaatt tcacacagga aacagctatg accatgatta cgccaagcgc
      481 gcaattaacc ctcactaaag ggaacaaaag ctggtctagc ttaagagttc accgacaaac
      541 aacagataaa acgaaaggcc cagtctttcg actgagcctt tcgttttatt tgatgcctgg
      601 cagttcccta ctctcgcatg gggagacccc acactaccat cggcgctacg gcgtttcact
      661 tctgagttcg gcatggggtc aggtggggcc accgcgctac tgccgccagg caaattctgt
      721 tttatcagac cgcttctgcg ttctgattta atctgtatca ggctgaaaat cttctctcat
      781 ccgccaaaac agccaagctg gagaccgttt aaactcaatg atgatgatga tgatggtcga
      841 cggcgctatt cagatcctct tctgagatga gtttttgttc gggcccaagc ttcaaaaaaa
      901 gcacatgggt gatcctcatg tgccactttt tcaagttgat aacggactag ccttatttta
      961 acttgctatt tctagctcta aaacacggag cgttctggac acaaactagt attataccta
     1021 ggactgagct agctgtcaag agctccaatt cgccctatag tgagtcgtat tacgcgcgct
     1081 cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc
     1141 gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc
     1201 gcccttccca acagttgcgc agcctgaatg gcgaatggaa attgtaagcg ttaatatttt
     1261 gttaaaattc gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgtact
     1321 gcgatgagtg gcagggcggg gcgtaatttt tttaaggcag ttattggtgc ccttaaacgc
     1381 ctggttgcta cgcctgaata agtgataata agcggatgaa tggcagaaat tcgaaagcaa
     1441 attcgacccg gtcgtcggtt cagggcaggg tcgttaaata gccgcttatg tctattgctg
     1501 gtttaccggt ttattgacta ccggaagcag tgtgaccgtg tgcttctcaa atgcctgagg
     1561 ccagtttgct caggctctcc ccgtggaggt aataattgac gatatgatca tttattctgc
     1621 ctcccagagc ctgataaaaa cggtgaatcc gttagcgagg tgccgccggc ttccattcag
     1681 gtcgaggtgg cccggctcca tgcaccgcga cgcaacgcgg ggaggcagac aaggtatagg
     1741 gcggcgaggc ggctacagcc gatagtctgg aacagcgcac ttacgggttg ctgcgcaacc
     1801 caagtgctac cggcgcggca gcgtgacccg tgtcggcggc tccaacggct cgccatcgtc
     1861 cagaaaacac ggctcatcgg gcatcggcag gcgctgctgc ccgcgccgtt cccattcctc
     1921 cgtttcggtc aaggctggca ggtctggttc catgcccgga atgccgggct ggctgggcgg
     1981 ctcctcgccg gggccggtcg gtagttgctg ctcgcccgga tacagggtcg ggatgcggcg
     2041 caggtcgcca tgccccaaca gcgattcgtc ctggtcgtcg tgatcaacca ccacggcggc
     2101 actgaacacc gacaggcgca actggtcgcg gggctggccc cacgccacgc ggtcattgac
     2161 cacgtaggcc gacacggtgc cggggccgtt gagcttcacg acggagatcc agcgctcggc
     2221 caccaagtcc ttgactgcgt attggaccgt ccgcaaagaa cgtccgatga gcttggaaag
     2281 tgtcttctgg ctgaccacca cggcgttctg gtggcccatc tgcgccacga ggtgatgcag
     2341 cagcattgcc gccgtgggtt tcctcgcaat aagcccggcc cacgcctcat gcgctttgcg
     2401 ttccgtttgc acccagtgac cgggcttgtt cttggcttga atgccgattt ctctggactg
     2461 cgtggccatg cttatctcca tgcggtaggg gtgccgcacg gttgcggcac catgcgcaat
     2521 cagctgcaac ttttcggcag cgcgacaaca attatgcgtt gcgtaaaagt ggcagtcaat
     2581 tacagatttt ctttaaccta cgcaatgagc tattgcgggg ggtgccgcaa tgagctgttg
     2641 cgtacccccc ttttttaagt tgttgatttt taagtctttc gcatttcgcc ctatatctag
     2701 ttctttggtg cccaaagaag ggcacccctg cggggttccc ccacgccttc ggcgcggctc
     2761 cccctccggc aaaaagtggc ccctccgggg cttgttgatc gactgcgcgg ccttcggcct
     2821 tgcccaaggt ggcgctgccc ccttggaacc cccgcactcg ccgccgtgag gctcgggggg
     2881 caggcgggcg ggcttcgccc ttcgactgcc cccactcgca taggcttggg tcgttccagg
     2941 cgcgtcaagg ccaagccgct gcgcggtcgc tgcgcgagcc ttgacccgcc ttccacttgg
     3001 tgtccaaccg gcaagcgaag cgcgcaggcc gcaggccgga ggcttttccc cagagaaaat
     3061 taaaaaaatt gatggggcaa ggccgcaggc cgcgcagttg gagccggtgg gtatgtggtc
     3121 gaaggctggg tagccggtgg gcaatccctg tggtcaagct cgtgggcagg cgcagcctgt
     3181 ccatcagctt gtccagcagg gttgtccacg ggccgagcga agcgagccag ccggtggccg
     3241 ctcgcggcca tcgtccacat atccacgggc tggcaaggga gcgcagcgac cgcgcagggc
     3301 gaagcccgga gagcaagccc gtagggcgcc gcagccgccg taggcggtca cgactttgcg
     3361 aagcaaagtc tagtgagtat actcaagcat tgagtggccc gccggaggca ccgccttgcg
     3421 ctgcccccgt cgagccggtt ggacaccaaa agggaggggc aggcatggcg gcatacgcga
     3481 tcatgcgatg caagaagctg gcgaaaatgg gcaacgtggc ggccagtctc aagcacgcct
     3541 accgcgagcg cgagacgccc aacgctgacg ccagcaggac gccagagaac gagcactggg
     3601 cggccagcag caccgatgaa gcgatgggcc gactgcgcga gttgctgcca gagaagcggc
     3661 gcaaggacgc tgtgttggcg gtcgagtacg tcatgacggc cagcccggaa tggtggaagt
     3721 cggccagcca agaacagcag gcggcgttct tcgagaaggc gcacaagtgg ctggcggaca
     3781 agtacggggc ggatcgcatc gtgacggcca gcatccaccg tgacgaaacc agcccgcaca
     3841 tgaccgcgtt cgtggtgccg ctgacgcagg acggcaggct gtcggccaag gagttcatcg
     3901 gcaacaaagc gcagatgacc cgcgaccaga ccacgtttgc ggccgctgtg gccgatctag
     3961 ggctgcaacg gggcatcgag ggcagcaagg cacgtcacac gcgcattcag gcgttctacg
     4021 aggccctgga gcggccacca gtgggccacg tcaccatcag cccgcaagcg gtcgagccac
     4081 gcgcctatgc accgcaggga ttggccgaaa agctgggaat ctcaaagcgc gttgagacgc
     4141 cggaagccgt ggccgaccgg ctgacaaaag cggttcggca ggggtatgag cctgccctac
     4201 aggccgccgc aggagcgcgt gagatgcgca agaaggccga tcaagcccaa gagacggccc
     4261 gagaccttcg ggagcgcctg aagcccgttc tggacgccct ggggccgttg aatcgggata
     4321 tgcaggccaa ggccgccgcg atcatcaagg ccgtgggcga aaagctgctg acggaacagc
     4381 gggaagtcca gcgccagaaa caggcccagc gccagcagga acgcgggcgc gcacatttcc
     4441 ccgaaaagtg ccacctgcgg cgttgtgaca atttaccgaa caactccgcg gccgggaagc
     4501 cgatctcggc ttgaacgaat tgttaggtgg cggtacttgg gtcgatatca aagtgcatca
     4561 cttcttcccg tatgcccaac tttgtataga gagccactgc gggatcgtca ccgtaatctg
     4621 cttgcacgta gatcacataa gcaccaagcg cgttggcctc atgcttgagg agattgatga
     4681 gcgcggtggc aatgccctgc ctccggtgct ctccggagac tgcgagatca tagatataga
     4741 tctcactacg cggctgctca aacttgggca gaacgtaagc cgcgagagcg ccaacaaccg
     4801 cttcttggtc gaaggcagca agcgcgatga atgtcttact acggagcaag ttcccgaggt
     4861 aatcggagtc cggctgatgt tgggagtagg tggctacgtc tccgaactca cgaccgaaaa
     4921 gatcaagagc agcccgcatg gatttgactt ggtcagggcc gagcctacat gtgcgaatga
     4981 tgcccatact tgagccacct aactttgttt tagggcgact gccctgctgc gtaacatcgt
     5041 tgctgctgcg taacatcgtt gctgctccat aacatcaaac atcgacccac ggcgtaacgc
     5101 gcttgctgct tggatgcccg aggcatagac tgtacaaaaa aacagtcata acaagccatg
     5161 aaaaccgcca ctgcgccgtt accaccgctg cgttcggtca aggttctgga ccagttgcgt
     5221 gagcgcatac gctacttgca ttacagttta cgaaccgaac aggcttatgt caactgggtt
     5281 cgtgccttca tccgtttcca cggtgtgcgt ccatgggcaa atattatacg caaggcgaca
     5341 aggtgctgat gccgctggcg attcaggttc atcatgccgt ttgtgatggc ttccatgtcg
     5401 gcagaatgct taatgaatta caacagtttt tatataaacg cagaaaggcc cacccgaagg
     5461 tgagccagtg tgactctagt agagagcgtt caccgacaaa caacagataa aacgaaaggc
     5521 ccagtctttc gactgagcct ttcgttttat ttgatgcctg gagatcctta ctcgagtttt
     5581 tattcgtcgt agaaatcaat gtggccaccg gaggtgaagg tgtcagcctt taacaaatga
     5641 atcaactttc tagaggagtc ctccatcttc accaaggaac cagacttatg catgtcgtcg
     5701 tagatcttca gttgttcctt atcacccaaa gttctgcgga cgtcagcctg catttcgtta
     5761 tccaatggac ctggagcgta gttcaaggtc tttacgttat tagctgcttc ttcgagagca
     5821 ataacaccta atagtctatc tctagctgcc ttaccagcag cgtacaaacc ccagtttggg
     5881 aaagcctgaa cagctaatag ggaagagatg gacacgacca cagttctatg atctgggtat
     5941 tgttcctttg ggaatgcttc gagggtatcc ttcaagaaca tggagcacaa accaacgagg
     6001 gacacgacgt tgaaatcgag gtaggaacga gcctcttgcc aggtgaactc cttaacagtc
     6061 ttggacaagt cacccaagct acccgcgttg ttgaagagga ccgacttagt gatagttctg
     6121 gtgctttgcg ccgcctcgtc tcttaggtca gcggcggcgg actgtatcct cgcgaggttg
     6181 gagtcgaggg agttcaagtg agccaggtcg acctcggaca caacgacacc cttgaagacg
     6241 acaccagcgt gggatgcagc ctcgtgcagc tccgttaaga cttgctccaa ggaatgctgg
     6301 gttctaccaa ccaagacaaa ggacaccgcg tgagcacctg agtgtcttat gtaatccaac
     6361 gcgacggagt gcccgaaacc tctattaaca ccgttaataa taaccaaatg atgttcttta
     6421 ctgctatggt ggtgatggtg atgcatggta cctttctcct ctttaatgaa ttcgctagca
     6481 caatccctag gactgagcta gctgtcaatc tataatcgca acttcaagac gacgttgtgt
     6541 ccagaacgct ccgtaggaga agtgaggaga cgagcgaacg cgtcgtacga gctttatgca
     6601 tctttcacag ctcgtagcga tgattatgac atcgcagtga gcaactgaat aagcgctgcg
     6661 tgaatgatcg caaatgctga ttataaacgc agaaaggccc acccgaaggt gagccagtgt
     6721 gactctagta gagagcgttc accgacaaac aacagataaa acgaaaggcc cagtctttcg
     6781 actgagcctt tcgttttatt tgatgcctgg ttattcacct ctgtaaacga caacgttctt
     6841 gtcggtttcg tgcaacttga tctcgtacaa atcgtatgcg tcggaaggtg gcaagtggct
     6901 cttgatattc tcccacaaga agacagcgag gttctcagta gtggagggtc tggactcgaa
     6961 gtatgggacg tctatatcca agtttctatg gtcacaaggg tccatgacag cgacttgcaa
     7021 agtcttctta agatcggtga tgttgatgac catgccggat tgtgggttga tctgaccctt
     7081 gatggtcacc tcgaccttgt agttgtgacc gtgaccggaa gtgtggttgc acttaccgaa
     7141 gagcttgacg ttctcagcag gcgagaggtg gacggagttc aatctgtgcg cagcggagaa
     7201 gtgttcgatc ctggtaacgt aagcagttct aactggagtt gaggacgtat ggtgatggtg
     7261 gtgatgcatg gtacctttct cctctttaat gaattcgcta gcacaatccc taggactgag
     7321 ctagctgtca atctataatc gcaacttcaa gacgacgttg tgtccagaac gctccgtagg
     7381 agaagtgagg agacgagcga acgcgtcgta cgagctttat gcatctttca cagctcgtag
     7441 cgatgattat gacatcgcag tgagcaactg aataagcgct gcgtgaatga tcgcaaatgc
     7501 tgattataaa cgcagaaagg cccacccgaa ggtgagccag tgtgactcta gtagagagcg
     7561 ttcaccgaca aacaacagat aaaacgaaag gcccagtctt tcgactgagc ctttcgtttt
     7621 atttgatgcc tggttagttg tgatgacgca cagcgcgcag aaactcgtgg cgcgtattct
     7681 gactggattt gaacaatcca ccaagagagg tcgttgtcgt ggcactggtt gcatcgcgga
     7741 tgccacgcgc cttcacgcag taatgcaccg cgtcgatcga gacagccacg ttattggtgc
     7801 ccagcagcgt ttgtagcgca ataagaattt gctgcgtcag acgttcctgc acctgcggac
     7861 gctgggcaaa gaactgcaca atgcggttaa tttttgacag accgatcacc gaatctttcg
     7921 ggatataggc caccgtcgct ttgccatcga tggtaacaaa atggtgttca caggtgctgg
     7981 tcagagtgat atcgcgcacg gtgaccattt catcgacctt cattttgttt tcaatgaggg
     8041 tgattttcgg gaaattggcg taatccagac cggagaaaat ttcatcgaca tacattttag
     8101 cgatgcgatg cggcgtttcc atcaaactgt catcagccag gtcgagattc agcagctgca
     8161 tgatttcggt catatgacca gcaataaggc ttttgcgcgt ttcgttatcc atttcatgca
     8221 cgggcgggcg cagcggtgtt tccagtcctc gcgcaactaa cgcttcatga accagggccg
     8281 cttctttact gagtgatggg tgatggtgat ggtgatgcat ggtacct
//
