LOCUS       sequence-413680-       10252 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Production of syn-pimara-7,15-diene in E. coli MEV20.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10252)
  AUTHORS   Lin GM, Voigt CA
  TITLE     Design of a redox-proficient Escherichia coli for screening 
            terpenoids and modifying cytochrome P450s
  JOURNAL   Nature Catalysis. 6, 1016–1029 (2023). 
            https://doi.org/10.1038/s41929-023-01049-5
REFERENCE   2  (bases 1 to 10252)
  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: "Nature 
            Catalysis. 6, 1016–1029 (2023). 
            https://doi.org/10.1038/s41929-023-01049-5"
COMMENT     Sequence Label: pLRD14
FEATURES             Location/Qualifiers
     source          1..10252
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     68..86
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     terminator      complement(90..133)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     misc_feature    136..157
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        178..196
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    197..221
                     /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)."
     promoter        1752..1770
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1771..1795
                     /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)."
     promoter        4188..4206
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    4207..4231
                     /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)."
     misc_feature    6800..6820
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      6821..6878
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     primer_bind     complement(6902..6919)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      7453..7675
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             7723..8673
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFWIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     CDS             complement(join(9287..10078,10079..10147))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     9910..9929
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10148..10252)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cacatttccc cgaaaagtgc cacctgacgt ctaagaaacc attattatca tgacattaac
       61 ctataaaaat aggcgtatca cgaggcagaa tttcagataa aaaaaatcct tagctttcgc
      121 taaggatgat ttctggaatt cgcggccgct tctagagcgc gaagacttaa tcaaatctaa
      181 tacgactcac tataggggaa ttgtgagcgg ataacaattc ccctctacgt tttcaaggtg
      241 gtagactgtc gccggatgtg tatccgacct gacgatggcc caaaagggcc gaaacagtcc
      301 tctacaaata attttgttta agagaatacc ccgctaagaa ggtaattatg tccatgcaaa
      361 gtttagataa gaattttcga catttatctc gtaaagaaaa gttacaacaa ttggttgata
      421 agcaatggtt atcagaagaa caattcgaca ttttactgaa tcatccatta atcgatgaag
      481 aagtagccaa tagtttaatt gaaaatgtca tcgcgcaagg tgcattaccc gttggattat
      541 taccgaatat cattgtggac gataaggcat atgttgtacc tatgatggtg gaagagcctt
      601 cagttgtcgc tgcagctagt tatggtgcaa agctagtgaa tcagactggc ggatttaaaa
      661 cggtatcttc tgaacgtatt atgataggtc aaatcgtctt tgatggcgtt gacgatactg
      721 aaaaattatc agcagacatt aaagctttag aaaagcaaat tcataaaatt gcggatgagg
      781 catatccttc tattaaagcg cgtggtggtg gttaccaacg tatagcgatt gatacatttc
      841 ctgagcaaca gttactatct ttaaaagtat ttgttgatac gaaagatgct atgggcgcta
      901 atatgcttaa tacgatttta gaggccataa ctgcattttt aaaaaatgaa tttccgcaaa
      961 gcgacatttt aatgagtatt ttatccaatc atgcaacagc gtccgttgtt aaagttcaag
     1021 gcgaaattga tgttaaagat ttagcaaggg gcgagagaac tggagaagag gttgccaaac
     1081 gaatggaacg tgcttctgta ttggcccaag tagatattca tcgtgcagca acacataata
     1141 aaggtgttat gaatggcata catgctgttg ttttagcaac aggaaatgat acgcgtggtg
     1201 cagaagcaag tgcgcatgca tacgcgagtc gtgacggaca gtatcgtggt attgctacat
     1261 ggcgttacga tcaagatcgt caacgattga ttggtacaat tgaagtgcct atgacattgg
     1321 caatcgttgg cggtggtaca aaagtattac caattgctaa agcttcatta gagctactaa
     1381 atgtagagtc agcacaagaa ttaggtcatg tagttgctgc cgttggttta gcgcaaaact
     1441 ttgcagcatg tcgcgcgctt gtgtcagaag gtattcaaca aggtcatatg agtttacaat
     1501 ataaatcatt agctatcgtt gtaggggcaa aaggtgatga aattgctaaa gtagctgaag
     1561 ctttgaaaaa agaaccccgt gcaaatacac aagcagcgga acatatttta caagaaatta
     1621 gacaacaata ataacggatg taaacagaga aacgtctaac taaaaaggcc tcccaaatcg
     1681 gggggccttt tttattgata acaaaagcct aatacacctg ttcgtgacga ctacggatgc
     1741 gggcaaggtg ctaatacgac tcactatagg ggaattgtga gcggataaca attcccctct
     1801 aaacgccagc cactaaagcg gtcaacgcat gtgctttgcg ttctgatgag acagtgatgt
     1861 cgaaaccgcc tctacaaata attttgttta agaacctaat tttataagga acacctaatg
     1921 ctgatttcaa aaagcccgcc ttaccctgcg tccgaagaaa cacgggaatg ggaggccgag
     1981 ggccagcatg aacataccga tgaattgcgt gaaacaacca cgacgatgat tgatggcatc
     2041 agaacagccc tgcgcagtat cggggaaggg gaaatctcga ttagcgccta cgatacgtca
     2101 ctggtagcgc tcctgaaacg cctggatggt ggggatgggc cgcagttccc gtcaacgatc
     2161 gattggatcg tacagaatca gttgcccgat ggtagctggg gcgatgcgag cttctttatg
     2221 atgggagatc gcatcatgtc caccctggcg tgcgtagttg ctctcaaatc ttggaacatt
     2281 cataccgaca aatgtgaacg tggtctgctg ttcattcagg agaatatgtg gcggttggct
     2341 cacgaagaag aagattggat gcttgtgggg ttcgaaatcg cgctcccatc gctgctggac
     2401 atggccaagg atttggatct cgatattcca tacgatgagc ctgctctgaa ggctatttac
     2461 gcggaacgcg agcgtaaatt ggcgaaaatc ccacgtgacg tgttacacgc tatgcctacc
     2521 acgctgctgc actccctgga aggcatggtg gatctggatt gggaaaaact tctgaaactg
     2581 cgctgtttgg acggaagttt ccactgcagc cctgcatcga ccgccaccgc gtttcaacaa
     2641 accggcgacc agaaatgctt cgagtacctg gatggtattg tcaagaaatt caatggcggc
     2701 gttccgtgca tctatccgct tgatgtctat gaacgcctgt gggccgttga ccgcctcacc
     2761 cgtctgggta tttcgcgtca ttttacatcc gaaatcgagg attgcctgga ctacattttc
     2821 cgcaactgga cgcccgacgg cctggctcat accaaaaact gcccggtaaa agatatagat
     2881 gataccgcta tgggatttcg acttttacgc ctgtatggat accaggtaga tccctgcgtg
     2941 ctgaagaaat tcgaaaaaga tggcaaattt ttctgccttc atggcgaaag caacccgtca
     3001 tctgtaacac cgatgtataa cacgtaccgc gcaagccaac tgaaatttcc gggtgatgac
     3061 ggggtgttag gtcgtgcaga ggtcttttgt cgcagctttc tccaggatcg ccgaggctcc
     3121 aatcgtatga aagataaatg ggctatcgcg aaagacattc ccggcgaggt tgaatatgcc
     3181 atggattatc cttggaaggc gtccttaccg cgtatagaaa cccgcttgta tctggatcag
     3241 tatggtggta gcggcgatgt ctggattggc aaagttctgc accgtatgac gctgttttgc
     3301 aatgatctgt atttgaaagc ggcgaaggcg gacttctcga attttcagaa agaatgccgc
     3361 gtagagctta atggcctgcg tcgttggtat ctgcgttcca atctcgagcg ttttggcggc
     3421 acggacccgc aaacgaccct gatgacctct tattttcttg cgtccgccaa catattcgag
     3481 ccgaaccgcg ccgccgaacg tcttggttgg gcacgtgtag ccctgctggc cgatgccgtt
     3541 agttcgcact ttcggcgcat cggtgggccg aaaaatctga ccagtaacct tgaggaatta
     3601 atctccctgg ttccctttga tgatgcctat tccggctccc tgcgagaagc gtggaagcag
     3661 tggttgatgg cctggacggc gaaagaatcg tcccaggaaa gtatcgaggg ggataccgcc
     3721 atattgttgg tgcgagcgat tgagatcttt gggggacgcc atgtgctgac tggtcagcgt
     3781 ccggaccttt gggaatattc tcagctggaa caattaacca gcagcatttg ccgcaaactt
     3841 tatagacggg tgctggcaca agaaaatggc aagagcactg agaaagtaga agagattgac
     3901 cagcagttag atctcgagat gcaggagttg acccgccgcg tgctgcaggg ctgcagcgct
     3961 attaaccgtc ttacacgtga gacattcctt catgtggtga agtctttctg ttacgttgca
     4021 tattgcagcc ccgaaaccat cgacaaccac attgacaaag ttatcttcca ggatgtcatc
     4081 taataattat gccttccgct aatcaactcg gtaccaaatt ccagaaaaga gacgctttcg
     4141 agcgtctttt ttcgttttgg tccgtggaca agcacagcat agccaagtaa tacgactcac
     4201 tataggggaa ttgtgagcgg ataacaattc ccctctaggt ccgaagatac ttagggtgtc
     4261 tcaaggtgcg taccttgact gatgagtccg aaaggacgaa acacccctct acaaataatt
     4321 ttgtttaaga ttagaatacc agaaagaagg tttttttatg gccgttatga gctcgtgccc
     4381 tggctacgtg gaagggcgaa tggtgggcga aaatacctct caaataaaca tggggcgtga
     4441 agcgcggatt cggagacact tggagaatcc ggagttcctg ccttctagct atgatatcgc
     4501 ttgggttgca atggtcccgc tcccaggcac ggatcatctt caggccccat gcttccccga
     4561 atgcgtcgaa tggatactgc aaaaccaaca tagcaatggc tcgtggggag ttaatgaatt
     4621 cgactcgtct gccagcaaag atattctgct gagtaccctt gcatgtatta ttgcccttga
     4681 aaaatggaat gtaggtagtg aacaaattcg ccgcggcctg cacttcatcg cgaagaattt
     4741 ttctattgtg atcgacgatc agatcgctgc gccgattggc tttaacctga ccttcccggc
     4801 aatggtaaac ttggcaatta aaatgggcct tgaatttcct gcgtcggaaa ttagcatcga
     4861 ccagatcctg catctccgtg acatggagct caagcgcctg tccggagaag aatctctggg
     4921 taaggaggcg tactttgctt acattgctga aggccttgaa gagagcatgg tcgactggtc
     4981 agaagtcatg aagttccagg gtaaaaacgg cagtctgttt aactcgcccg ccgcaacggc
     5041 cgcagctctg gtgcacagat atgatgacaa agccctgggt tacctctatt cggtggtcaa
     5101 caaatttggt ggtgaggtcc cgacggtgta tccccttaat atttttagtc aactgtcgat
     5161 ggtggatacg ttagtgaata tcggtatcag ccggcatttt tcttcagaca tcaaacgcat
     5221 tctcgataaa acctacatcc tgtggtccca acgtgatgag gaggttatgc tcgatcttcc
     5281 gacgtgcgca atggcgttcc gtctgctcag aatgaatggc tatggcgtta gtagcgatga
     5341 tctcagtcac gtcgccgaag cgtccacgtt tcataactca gttgagggct atctggatga
     5401 caccaaatcg ctgttggaac tgtataaagc atccaaagtg agcttatccg agaacgagcc
     5461 gattcttgag aaaatgggtt gctggtccgg cagtctcctc aaagaaaaac tgtgcagcga
     5521 cgatattcgc ggcaccccga ttttaggaga ggtagagtac gcccttaaat ttccatttta
     5581 tgctaccctg gaaccattag accacaaatg gaatattgag aattttgacg ccagagccta
     5641 tcagaagatt aaaaccaaaa acatgccctg ccatgtgaat gaggacctgt tagcgcttgc
     5701 ggctgaggat ttcagttttt gccagtctac gtaccaaaat gaaattcagc atctggaaag
     5761 ctgggaaaaa gagaacaaac tcgatcaact cgagttcact cgcaaaaatc tcataaactc
     5821 ctacttatcg gcggcggcca ctatttcccc gtatgaactg tcggatgcgc gcattgcatg
     5881 cgccaaaagc attgccttaa ccctcgtcgc cgatgatttc ttcgatgttg gatcttcaaa
     5941 ggaggaacag gaaaatctga tctccctggt ggaaaaatgg gatcagtatc acaaagttga
     6001 attctactct gagaatgtga aagccgtgtt ctttgccctg tattccacgg tgaaccagtt
     6061 gggggccatg gcgtctgcgg tgcagaatcg tgacgttacc aagtataacg ttgaaagctg
     6121 gcttgactat ttacgcagcc tggcaaccga cgcggagtgg caacgtagca aatacgttcc
     6181 aacaatggaa gaatacatga aaaattcgat agtgaccttt gccttaggcc cgaccatctt
     6241 gatcgcgctg tatttcatgg gtcagaatct gtgggaggac atcgtgaaaa atgcggaata
     6301 tgacgaactt tttcgcctca tgaacacatg tggccgttta caaaacgaca ttcagtcttt
     6361 cgaacgtgaa tgtaaagatg gtaaactgaa ttcggtttcc ttactggtac ttgattcgaa
     6421 ggatgtgatg tctgtggaag aggcaaagga agctatcaac gaatcgataa gctcctgccg
     6481 acgcgagttg ttgagactgg tggtccgcga ggacggggtc attcctaaga gctgtaagga
     6541 aatgttctgg aacctgtata aaacctccca tgtgttttac agtcaggccg atggcttcag
     6601 cagtccgaaa gagatgatgg gcgcgatgaa tggtgttatc ttcgaacccc tgaaaacccg
     6661 tggtaactga taagcctcgt tgctcttcat aaatcatccc tatgacccct tcaaaaaaaa
     6721 aaaacaccct aacgggtgtt tttttttttt tggtctgcct ctcgctgacg gtcacaatgc
     6781 gcttacgaaa gtcttcctgt actagtagcg gccgctgcag tccggcaaaa aagggcaagg
     6841 tgtcaccacc ctgccctttt tctttaaaac cgaaaagatt acttcgcgtt atgcaggctt
     6901 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     6961 caaaggcggt aatgacagta agacgggtaa gcctgttgat gataccgctg ccttactggg
     7021 tgcattagcc agtctgaatg acctgtcacg ggataatccg aagtggtcag actggaaaat
     7081 cagagggcag gaactgctga acagcaaaaa gtcagatagc accacatagc agacccgcca
     7141 taaaacgccc tgagaagccc gtgacgggct tttcttgtat tatgggtagt ttccttgcat
     7201 gaatccataa aaggcgcctg tagtgccatt tacccccatt cactgccaga gccgtgagcg
     7261 cagcgaactg aatgtcacga aaaagacagc gactcaggtg cctgatggtc ggagacaaaa
     7321 ggaatattca gcgatttgcc cgagcttgcg agggtgctac ttaagccttt agggttttaa
     7381 ggtctgtttt gtagaggagc aaacagcgtt tgcgacatcc ttttgtaata ctgcggaact
     7441 gactaaagta gtgagttata cacagggctg ggatctattc tttttatctt tttttattct
     7501 ttctttattc tataaattat aaccacttga atataaacaa aaaaaacaca caaaggtcta
     7561 gcggaattta cagagggtct agcagaattt acaagttttc cagcaaaggt ctagcagaat
     7621 ttacagatac ccacaactca aaggaaaagg actagtaatt atcattgact agcccatctc
     7681 aattggtata gtgattaaaa tcacctagac caattgagat gtatgtctga attagttgtt
     7741 ttcaaagcaa atgaactagc gattagtcgc tatgacttaa cggagcatga aaccaagcta
     7801 attttatgct gtgtggcact actcaacccc acgattgaaa accctacaag gaaagaacgg
     7861 acggtatcgt tcacttataa ccaatacgct cagatgatga acatcagtag ggaaaatgct
     7921 tatggtgtat tagctaaagc aaccagagag ctgatgacga gaactgtgga aatcaggaat
     7981 cctttggtta aaggcttttg gattttccag tggacaaact atgccaagtt ctcaagcgaa
     8041 aaattagaat tagtttttag tgaagagata ttgccttatc ttttccagtt aaaaaaattc
     8101 ataaaatata atctggaaca tgttaagtct tttgaaaaca aatactctat gaggatttat
     8161 gagtggttat taaaagaact aacacaaaag aaaactcaca aggcaaatat agagattagc
     8221 cttgatgaat ttaagttcat gttaatgctt gaaaataact accatgagtt taaaaggctt
     8281 aaccaatggg ttttgaaacc aataagtaaa gatttaaaca cttacagcaa tatgaaattg
     8341 gtggttgata agcgaggccg cccgactgat acgttgattt tccaagttga actagataga
     8401 caaatggatc tcgtaaccga acttgagaac aaccagataa aaatgaatgg tgacaaaata
     8461 ccaacaacca ttacatcaga ttcctaccta cataacggac taagaaaaac actacacgat
     8521 gctttaactg caaaaattca gctcaccagt tttgaggcaa aatttttgag tgacatgcaa
     8581 agtaagtatg atctcaatgg ttcgttctca tggctcacgc aaaaacaacg aaccacacta
     8641 gagaacatac tggctaaata cggaaggatc tgaggttctt atggctcttg tatctatcag
     8701 tgaagcatca agactaacaa acaaaagtag aacaactgtt caccgttaca tatcaaaggg
     8761 aaaactgtcc atatgcacag atgaaaacgg tgtaaaaaag atagatacat cagagctttt
     8821 acgagttttt ggtgcattca aagctgttca ccatgaacag atcgacaatg taacagatga
     8881 acagcatgta acacctaata gaacaggtga aaccagtaaa acaaagcaac tagaacatga
     8941 aattgaacac ctgagacaac ttgttacagc tcaacagtca cacatagaca gcctgaaaca
     9001 ggcgatgctg cttatcgaat caaagctgcc gacaacacgg gagccagtga cgcctcccgt
     9061 ggggaaaaaa tcatggcaat tctggaagaa atagcgcttt cagccggcaa accggctgaa
     9121 gccggatctg cgattctgat aacaaactag caacaccaga acagcccgtt tgcgggcagc
     9181 aaaacccgta ccgattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     9241 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     9301 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     9361 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     9421 taccgcgaga accacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     9481 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     9541 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     9601 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     9661 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     9721 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     9781 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     9841 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     9901 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     9961 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatataac
    10021 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
    10081 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
    10141 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
    10201 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cg
//
