LOCUS       sequence-327149-        8675 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Integrative expression of mRFP-CUP1-AID* (mini auxin inducible 
            degron)-yEGFP and rice auxin receptor gene OsTIR1. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8675)
  AUTHORS   Lu Z, Peng B, Ebert BE, Dumsday G, Vickers CE
  TITLE     Auxin-mediated protein depletion for metabolic engineering in 
            terpene-producing yeast.
  JOURNAL   Nat Commun. 2021 Feb 16;12(1):1051. doi: 10.1038/s41467-021-21313-1.
  PUBMED    33594068
REFERENCE   2  (bases 1 to 8675)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2021-02-16"; volume: "12(1):1051. doi"; pages: " 
            10.1038/s41467-021-21313-1"
COMMENT     Sequence Label: pJAID5XY
FEATURES             Location/Qualifiers
     source          1..8675
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             431..1105
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     CDS             1247..1957
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKVN
                     FKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     terminator      1973..2243
                     /gene="S. cerevisae PGK1"
                     /label=PGK1 terminator
                     /note="transcription terminator for 3-phosphoglycerate 
                     kinase gene"
     CDS             2739..4466
                     /codon_start=1
                     /product="rice transport inhibitor response 1 protein"
                     /label=OsTIR1
                     /note="F-box protein that mediates auxin-dependent 
                     interaction of a ubiquitin ligase with an auxin-inducible 
                     degron (Tan et al., 2007)"
                     /translation="MTYFPEEVVEHIFSFLPAQRDRNTVSLVCKVWYEIERLSRRGVFV
                     GNCYAVRAGRVAARFPNVRALTVKGKPHFADFNLVPPDWGGYAGPWIEAAARGCHGLEE
                     LRMKRMVVSDESLELLARSFPRFRALVLISCEGFSTDGLAAVASHCKLLRELDLQENEV
                     EDRGPRWLSCFPDSCTSLVSLNFACIKGEVNAGSLERLVSRSPNLRSLRLNRSVSVDTL
                     AKILLRTPNLEDLGTGNLTDDFQTESYFKLTSALEKCKMLRSLSGFWDASPVCLSFIYP
                     LCAQLTGLNLSYAPTLDASDLTKMISRCVKLQRLWVLDCISDKGLQVVASSCKDLQELR
                     VFPSDFYVAGYSAVTEEGLVAVSLGCPKLNSLLYFCHQMTNAALVTVAKNCPNFTRFRL
                     CILEPGKPDVVTSQPLDEGFGAIVRECKGLQRLSISGLLTDKVFMYIGKYAKQLEMLSI
                     AFAGDSDKGMMHVMNGCKNLRKLEIRDSPFGDAALLGNFARYETMRSLWMSSCNVTLKG
                     CQVLASKMPMLNVEVINERDGSNEMEENHGDLPKVEKLYVYRTTAGARDDAPNFVKIL"
     primer_bind     complement(4696..4712)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4696..4712)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4709..4731)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4720..4736
                     /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        complement(join(4744..4750,4751..4768,4769..4774))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4789..4810
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4927..4944)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5098..5686)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5178..5197)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5857..6648,6649..6717))
                     /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     6480..6499
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6718..6822)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6890..6908
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(6946..6968)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7068..7087
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7281..7303
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7295..7312
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     7296..7312
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(8426..8459)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        join(8670..8675,1..408)
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
ORIGIN
        1 ccacacacca tagcttcaaa atgtttctac tcctttttta ctcttccaga ttttctcgga
       61 ctccgcgcat cgccgtacca cttcaaaaca cccaagcaca gcatactaaa tttcccctct
      121 ttcttcctct agggtgtcgt taattacccg tactaaaggt ttggaaaaga aaaaagagac
      181 cgcctcgttt ctttttcttc gtcgaaaaag gcaataaaaa tttttatcac gtttcttttt
      241 cttgaaaatt tttttttttg atttttttct ctttcgatga cctcccattg atatttaagt
      301 taataaacgg tcttcaattt ctcaagtttc agtttcattt ttcttgttct attacaactt
      361 tttttacttc ttgctcatta gaaagaaagc atagcaatct aatctaagtt ttaattacaa
      421 actcgagaaa atggcttcct ccgaagacgt tatcaaagag ttcatgcgtt tcaaagttcg
      481 tatggaaggt tccgttaacg gtcacgagtt cgaaatcgaa ggtgaaggtg aaggtcgtcc
      541 gtacgaaggt acccagaccg ctaaactgaa agttaccaaa ggtggtccgc tgccgttcgc
      601 ttgggacatc ctgtccccgc agttccagta cggttccaaa gcttacgtta aacacccggc
      661 tgacatcccg gactacctga aactgtcctt cccggaaggt ttcaaatggg aacgtgttat
      721 gaacttcgaa gacggtggtg ttgttaccgt tacccaggac tcctccctgc aagacggtga
      781 gttcatctac aaagttaaac tgcgtggtac caacttcccg tccgacggtc cggttatgca
      841 gaaaaaaacc atgggttggg aagcttccac cgaacgtatg tacccggaag acggtgctct
      901 gaaaggtgaa atcaaaatgc gtctgaaact gaaagacggt ggtcactacg acgctgaagt
      961 taaaaccacc tacatggcta aaaaaccggt tcagctgccg ggtgcttaca aaaccgacat
     1021 caaactggac atcacctccc acaacgaaga ctacaccatc gttgaacagt acgaacgtgc
     1081 tgaaggtcgt cactccaccg gtgctccaaa agacccagct aaacctccag ccaaggccca
     1141 agtcgtcggc tggccaccag ttaggtctta cagaaagaac gtaatggtta gttgtcaaaa
     1201 gtcctccggt ggtccagaag ctgctgcctt tgttaagatg ggatcctcta aaggtgaaga
     1261 attattcact ggtgttgtcc caattttggt tgaattagat ggtgatgtta atggtcacaa
     1321 attttctgtc tccggtgaag gtgaaggtga tgctacttac ggtaaattga ccttaaaatt
     1381 tatttgtact actggtaaat tgccagttcc atggccaacc ttagtcacta ctttcggtta
     1441 tggtgttcaa tgttttgcga gatacccaga tcatatgaaa caacatgact ttttcaagtc
     1501 tgccatgcca gaaggttatg ttcaagaaag aactattttt ttcaaagatg acggtaacta
     1561 caagaccaga gctgaagtca agtttgaagg tgatacctta gttaatagaa tcgaattaaa
     1621 aggtattgat tttaaagaag atggtaacat tttaggtcac aaattggaat acaactataa
     1681 ctctcacaat gtttacatca tggctgacaa acaaaagaat ggtatcaaag ttaacttcaa
     1741 aattagacac aacattgaag atggttctgt tcaattagct gaccattatc aacaaaatac
     1801 tccaattggt gatggtccag tcttgttacc agacaaccat tacttatcca ctcaatctgc
     1861 cttatccaaa gatccaaacg aaaagagaga ccacatggtc ttgttagaat ttgttactgc
     1921 tgctggtatt acccatggta tggatgaatt gtacaaaaga tcttaaacta gtattgaatt
     1981 gaattgaaat cgatagatca atttttttct tttctctttc cccatccttt acgctaaaat
     2041 aatagtttat tttatttttt gaatattttt tatttatata cgtatatata gactattatt
     2101 tatcttttaa tgattattaa gatttttatt aaaaaaaaat tcgctcctct tttaatgcct
     2161 ttatgcagtt tttttttccc attcgatatt tctatgttcg ggttcagcgt attttaagtt
     2221 taataactcg aaaattctgc gttcgttaaa gctttcgaga aggatattat ttgcggccgc
     2281 gattgcaaga attgatctcc ctcatcgatg tcataaatca gtaaatataa acattaaata
     2341 cattacacgt agccacccgc tgtatagcat aaaactaact taaaattatt catatgcgtt
     2401 tcccggggcc gaagcgttat tgccgatatt cggtagccga ttccgctggc tttttttttt
     2461 tgaacgaggc aaaaaagttt cgtgactgaa cttcttcgtc actggttttg ttgataattt
     2521 ttatcgtttt ttacttcgag attgaaaaca gtgttctgaa ttttagcaac gatatcgcac
     2581 cgcacgttta gcctgatcaa accttatata tacctctacc ttaaaatata tatatatata
     2641 gtattcgaat tccgaattgt gtgtctccgc ggttagtgat tgttatacac aaacagaata
     2701 caggaaagta aatcaataca ataataaatt ctagcaaaat gacctatttc ccagaagaag
     2761 tcgttgaaca catcttctct ttcttgccag ctcaaagaga tagaaatact gtttccttgg
     2821 tttgtaaggt ttggtatgaa atcgaaagat tgagtagacg tggtgttttc gtcggtaatt
     2881 gttacgctgt tagagctggt agagttgctg ctagattccc aaatgttaga gccttgactg
     2941 tcaaaggtaa accacatttt gctgacttta acttggtccc accagattgg ggtggttatg
     3001 ctggtccatg gattgaagct gccgctagag gttgtcacgg tttggaagaa ttgcgtatga
     3061 aaagaatggt cgtttctgac gaatctttgg aattattggc ccgttctttt ccaagattca
     3121 gagctttggt tttgatctct tgtgaaggtt tcagtactga cggtttggct gccgttgctt
     3181 cccattgtaa attattgaga gaattggact tgcaagaaaa tgaagtcgaa gatagaggtc
     3241 caagatggtt atcttgtttt ccagactctt gtacttcttt ggtctctttg aacttcgctt
     3301 gtattaaggg tgaagtcaac gctggttctt tagaaagatt ggtctcccgt tctccaaact
     3361 tacgttcttt gagattgaat cgttctgttt ccgttgatac cttggctaaa atcttattga
     3421 gaaccccaaa cttggaagac ttaggtactg gtaacttgac cgacgatttc caaactgaat
     3481 cctactttaa attaacttct gctttagaaa agtgtaagat gttgagatcc ttatctggtt
     3541 tttgggacgc ctctccagtc tgtttatctt ttatctatcc attgtgtgct caattaactg
     3601 gtttaaactt atcttacgct ccaaccttgg atgctagtga cttaaccaag atgattagta
     3661 gatgtgttaa gttacaaaga ttatgggttt tagattgtat ttccgataag ggtttacaag
     3721 tcgttgctag tagctgtaag gatttgcaag aattaagagt ctttccaagt gatttttacg
     3781 ttgctggtta cagcgctgtt actgaagaag gtttggtcgc tgtttctttg ggttgtccaa
     3841 agttgaacag cttattgtac ttttgtcatc aaatgactaa tgctgcctta gtcactgttg
     3901 ctaagaactg tccaaacttc acccgtttca gattgtgtat tttggaacca ggtaaaccag
     3961 acgttgtcac ctcccaacca ttggatgaag gtttcggtgc tatcgttaga gaatgtaagg
     4021 gtttgcaaag attatctatc tctggtttgt taactgacaa agttttcatg tacatcggta
     4081 aatatgccaa acaattagaa atgttgagca ttgctttcgc tggtgactct gacaagggta
     4141 tgatgcacgt tatgaatggt tgtaagaatt tgagaaagtt ggaaattaga gattctccat
     4201 tcggtgacgc tgccttattg ggtaattttg ctagatacga aactatgaga agtttgtgga
     4261 tgagttcctg taacgttacc ttgaaaggtt gtcaagtctt agccagcaag atgccaatgt
     4321 tgaacgtcga agtcatcaac gaaagagacg gtagtaacga aatggaagaa aatcacggtg
     4381 acttaccaaa ggtagaaaag ttgtacgtat acagaactac tgctggtgct agagatgatg
     4441 ctccaaattt cgtaaagatt ttgtaaagga ctatttgcaa agggaaggga tgctaaggta
     4501 gagggtgaac gttacagaaa agcaggctgg gaagcatatt tgagaagatg cggccagcaa
     4561 aactaaaaaa ctgtattata agtaaatgca tgtatactaa actcacaaat tagagcttca
     4621 atttaattat atcagttatt acccgggaat ctcggtcgta atgatttaaa tgcatgcaag
     4681 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     4741 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     4801 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     4861 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     4921 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     4981 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     5041 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     5101 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     5161 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     5221 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     5281 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     5341 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     5401 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     5461 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     5521 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     5581 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     5641 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     5701 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     5761 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     5821 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     5881 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     5941 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     6001 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     6061 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     6121 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     6181 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     6241 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     6301 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     6361 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     6421 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     6481 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     6541 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     6601 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     6661 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     6721 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6781 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6841 gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa ataggcgtat
     6901 cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     6961 gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     7021 gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg catcagagca
     7081 gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg taaggagaaa
     7141 ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt
     7201 gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag
     7261 ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgaattcat
     7321 ttaaattttt ttgattcggt aatctccgaa cagaaggaag aacgaaggaa ggagcacaga
     7381 cttagattgg tatatatacg catatgtagt gttgaagaaa catgaaattg cccagtattc
     7441 ttaacccaac tgcacagaac aaaaacctgc aggaaacgaa gataaaacat gtccacaaaa
     7501 tcatatacca gtagagctga gactcatgca agtccggttg catcgaaact tttacgttta
     7561 atggatgaaa agaagaccaa tttgtgtgct tctcttgacg ttcgttcgac tgatgagcta
     7621 ttgaaacttg ttgaaacgtt gggtccatac atttgccttt tgaaaacaca cgttgatatc
     7681 ttggatgatt tcagttatga gggtactgtc gttccattga aagcattggc agagaaatac
     7741 aagttcttga tatttgagga cagaaaattc gccgatatcg gtaacacagt caaattacaa
     7801 tatacatcgg gcgtttaccg tatcgcagaa tggtctgata tcaccaacgc ccacggggtt
     7861 actggtgctg gtattgttgc tggcttgaaa caaggtgcgc aagaggtcac caaagaacca
     7921 aggggattat tgatgcttgc tgaattgtct tccaagggtt ctctagcaca cggtgaatat
     7981 actaagggta ccgttgatat tgcaaagagt gataaagatt tcgttattgg gttcattgct
     8041 cagaacgata tgggaggaag agaagaaggg tttgattggc taatcatgac cccaggtgta
     8101 ggtttagacg acaaaggcga tgcattgggt cagcagtaca gaaccgtcga cgaagttgta
     8161 agtggtggat cagatatcat cattgttggc agaggacttt tcgccaaggg tagagatcct
     8221 aaggttgaag gtgaaagata cagaaatgct ggatgggaag cgtaccaaaa gagaatcagc
     8281 gctccccatt aattatacag gaaacttaat agaacaaatc acatatttaa tctaatagcc
     8341 acctgcattg gcacggtgca acactacttc aacttcatct tacaaaaaga tcacgtgatc
     8401 tgttgtattg ggatctctag acctaataac ttcgtatagc atacattata cgaagttata
     8461 ttaagggttg ctcgaccaga aagcgaccac ccaactttgg ctgataatag cgtataaaca
     8521 atgcatactt tgtacgttca aaatacaatg cagtagatat atttatgcat attacatata
     8581 atacatatca cataggaagc aacaggcgcg ttggactttt aattttcgag gaccgcgaat
     8641 ccttacatca cacccaatcc cccacaagtg atccc
//
