LOCUS       sequence-245796-        8319 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses pTEF2-mKate2-tIDP1 and pTRX2_5xUAS-yEGFP-tCYC1 in yeast 
            from plasdmid pRS416.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8319)
  AUTHORS   Zhang J, Sonnenschein N, Pihl TP, Pedersen KR, Jensen MK, Keasling 
            JD
  TITLE     Engineering an NADPH/NADP(+) Redox Biosensor in Yeast.
  JOURNAL   ACS Synth Biol. 2016 Dec 16;5(12):1546-1556. doi: 
            10.1021/acssynbio.6b00135. Epub 2016 Jul 25.
  PUBMED    27419466
REFERENCE   2  (bases 1 to 8319)
  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"; doi: 
            "10.1021/acssynbio.6b00135"; journalName: "ACS Synth Biol"; date: 
            "2016-12-16- 16"; volume: "5"; issue: "12"; pages: "1546-1556"
COMMENT     Sequence Label: pCfB3236
FEATURES             Location/Qualifiers
     source          1..8319
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             211..1014
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(1145..1600)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1282..1303)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1494..1513
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1726..1748
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1740..1757
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1741..1757
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1767..1786
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1767..1785
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      complement(1814..1979)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     1900..1924
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             2895..3611
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(5347..5367)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5347..5365)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5386..5402)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5386..5402)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5399..5421)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5410..5426
                     /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(5434..5440,5441..5458,5459..5464))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5479..5500
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5617..5634)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5788..6376)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(join(6547..7338,7339..7407))
                     /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     7170..7189
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7408..7512)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    7549..8052
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     8093..8111
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(8149..8171)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     8271..8290
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        join(8314..8319,1..210)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
ORIGIN
        1 tcatcatttt ttttttattc ttttttttga tttcggtttc cttgaaattt ttttgattcg
       61 gtaatctccg aacagaagga agaacgaagg aaggagcaca gacttagatt ggtatatata
      121 cgcatatgta gtgttgaaga aacatgaaat tgcccagtat tcttaaccca actgcacaga
      181 acaaaaacct gcaggaaacg aagataaatc atgtcgaaag ctacatataa ggaacgtgct
      241 gctactcatc ctagtcctgt tgctgccaag ctatttaata tcatgcacga aaagcaaaca
      301 aacttgtgtg cttcattgga tgttcgtacc accaaggaat tactggagtt agttgaagca
      361 ttaggtccca aaatttgttt actaaaaaca catgtggata tcttgactga tttttccatg
      421 gagggcacag ttaagccgct aaaggcatta tccgccaagt acaatttttt actcttcgaa
      481 gacagaaaat ttgctgacat tggtaataca gtcaaattgc agtactctgc gggtgtatac
      541 agaatagcag aatgggcaga cattacgaat gcacacggtg tggtgggccc aggtattgtt
      601 agcggtttga agcaggcggc agaagaagta acaaaggaac ctagaggcct tttgatgtta
      661 gcagaattgt catgcaaggg ctccctatct actggagaat atactaaggg tactgttgac
      721 attgcgaaga gcgacaaaga ttttgttatc ggctttattg ctcaaagaga catgggtgga
      781 agagatgaag gttacgattg gttgattatg acacccggtg tgggtttaga tgacaaggga
      841 gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg tctctacagg atctgacatt
      901 attattgttg gaagaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt
      961 tacagaaaag caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact
     1021 gtattataag taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat
     1081 cagttattac cctatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     1141 caggaaattg taagcgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     1201 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     1261 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     1321 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     1381 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     1441 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     1501 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     1561 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt ccattcgcca ttcaggctgc
     1621 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggcgaaag
     1681 ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag tcacgacgtt
     1741 gtaaaacgac ggccagtgag cgcgcgtaat acgactcact atagggcgaa ttgggtaccg
     1801 ggccccccct cgagagcgac ctcatgctat acctgagaaa gcaacctgac ctacaggaaa
     1861 gagttactca agaataagaa ttttcgtttt aaaacctaag agtcacttta aaatttgtat
     1921 acacttattt tttttataac ttatttaata ataaaaatca taaatcataa gaaattcgct
     1981 tatttagaag tgtcaacaac gtatctacca acggaatgcg tgcgatcttg agtgccagtg
     2041 aatattaagt aatcatcaaa gtatatgtgt aattgtttat actcttagta aaggatgctc
     2101 cctacaaggt ggctcttttc ttactaagcg cgttcagttt ccagccagcc gaaagaggga
     2161 tatcagcttg agtgccagtg aatattaagt aatcatcaaa gtatatgtgt aattgtttat
     2221 actcttagta aaggatgctc cctacaaggt ggctcttttc ttactaagcg cgttcagttt
     2281 ccagccagcc gaaagaggga tatcagacct gcactcttga gtgccagtga atattaagta
     2341 atcatcaaag tatatgtgta attgtttata ctcttagtaa aggatgctcc ctacaaggtg
     2401 gctcttttct tactaagcgc gttcagtttc cagccagccg aaagagggat atcagcttga
     2461 gtgccagtga atattaagta atcatcaaag tatatgtgta attgtttata ctcttagtaa
     2521 aggatgctcc ctacaaggtg gctcttttct tactaagcgc gttcagtttc cagccagccg
     2581 aaagagggat atcagatctg tcatcttgag tgccagtgaa tattaagtaa tcatcaaagt
     2641 atatgtgtaa ttgtttatac tcttagtaaa ggatgctccc tacaaggtgg ctcttttctt
     2701 actaagcgcg ttcagtttcc agccagccga aagagggata tcagtatata agaaagccat
     2761 tcgggggatg aaaagctgac aagagaataa cgaggaccag tttttatttg ttgtctagca
     2821 agaattatac acgcacacat acacgagagt ctacgatatc tttaaataac acatcaataa
     2881 tctgtcataa aacaatgtct aaaggtgaag aattattcac tggtgttgtc ccaattttgg
     2941 ttgaattaga tggtgatgtt aatggtcaca aattttctgt ctccggtgaa ggtgaaggtg
     3001 atgctactta cggtaaattg accttaaaat ttatttgtac tactggtaaa ttgccagttc
     3061 catggccaac cttagtcact actttcggtt atggtgttca atgttttgcg agatacccag
     3121 atcatatgaa acaacatgac tttttcaagt ctgccatgcc agaaggttat gttcaagaaa
     3181 gaactatttt tttcaaagat gacggtaact acaagaccag agctgaagtc aagtttgaag
     3241 gtgatacctt agttaataga atcgaattaa aaggtattga ttttaaagaa gatggtaaca
     3301 ttttaggtca caaattggaa tacaactata actctcacaa tgtttacatc atggctgaca
     3361 aacaaaagaa tggtatcaaa gttaacttca aaattagaca caacattgaa gatggttctg
     3421 ttcaattagc tgaccattat caacaaaata ctccaattgg tgatggtcca gtcttgttac
     3481 cagacaacca ttacttatcc actcaatctg ccttatccaa agatccaaac gaaaagagag
     3541 accacatggt cttgttagaa tttgttactg ctgctggtat tacccatggt atggatgaat
     3601 tgtacaaata aatcgcgtgc attcatccgc tctaaccgaa aaggaaggag ttagacaacc
     3661 tgaagtctag gtccctattt atttttttat agttatgtta gtattaagaa cgttatttat
     3721 atttcaaatt tttctttttt ttctgtacag acgcgtgtac gcatgtaaca ttatactgaa
     3781 aaccttgctt gagaagagct cgagaacagc cccctaaaac acggtaatga agaagaagaa
     3841 ctcatgaaaa tgttggaaat tgatggtaat gatccgaact tggggaacct tgataaggaa
     3901 ttaaccgacg gttgcaaaca acaataaggt atatatattt atgataacag taaaaaaaaa
     3961 aaaaaaaaaa agtagttcat tatcctagag cataactaag tataacactt taaaaaatga
     4021 aaaaaaaaag tggtagattg ggctacgtaa attcgaagct ttatctgtgt cccaacttag
     4081 atggcaaatc acagtatcta gccactgcca cttcgtgttg ttcaacataa gtctctttat
     4141 cagcctcttt aatcctttct aatcttcggt cgacgtagta aacacctggc atctttaggt
     4201 ttttggctgg ctttttagaa cgataagtgg ttttcaaatt gcagattaga tgccctccac
     4261 caaccagttt aagggccatg tcagctctgc cttctaggcc accatcagca gggtacaggg
     4321 tttcggttga cgcttcccag cctaatgtct ttttctgcat aactggacca ttagaaggga
     4381 aattgacgcc tctaatcttg acattgtaga ttaaacaccc atcttgaagt gatgtgtctt
     4441 gagtggctgt caatacacct ccatcctcgt acgttgttac tctttcccat gtaaaaccct
     4501 ctggaaaact ctgtttgaaa aagtctggta taccttgagt atggtttatg aatgtcttgc
     4561 tcccatacat gaatgatgtt gccaagatat caaatgcaaa aggtaatggt ccaccttcta
     4621 cagctttaat tctcatagtt tgagttcctt cgtaaggttt gccttcaccc tcggatgtac
     4681 acttaaaatg atgattgttc acagtacctt ccatgtaaag tttcatgtgc atgttttcct
     4741 tgatgagttc agaaaccatg tttagttaat tatagttcgt tgaccgtata ttctaaaaac
     4801 aagtactcct taaaaaaaaa ccttgaaggg aataaacaag tagaatagat agagagaaaa
     4861 atagaaaatg caagagaatt tatatattag aaagagagaa agaaaaatgg aaaaaaaaaa
     4921 ataggaaaag ccagaaatag cactagaagg agcgacacca gaaaagaagg tgatggaacc
     4981 aatttagcta tatatagtta actaccggct cgatcatctc tgcctccagc atagtcgaag
     5041 aagaattttt tttttcttga ggcttctgtc agcaactcgt attttttctt tcttttttgg
     5101 tgagcctaaa aagttcccac gttctcttgt acgacgccgt cacaaacaac cttatgggta
     5161 atttgtcgcg gtctgggtgt ataaatgtgt gggtgcaaca tgaatgtacg gaggtagttt
     5221 gctgattggc ggtctataga taccttggtt atggcgccct cacagccggc aggggaagcg
     5281 cctacgcttg acatctacta tatgtaagta tacggcccca tatatctcga gctccagctt
     5341 ttgttccctt tagtgagggt taattgcgcg cttggcgtaa tcatggtcat agctgtttcc
     5401 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     5461 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     5521 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5581 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     5641 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     5701 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     5761 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     5821 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     5881 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     5941 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     6001 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     6061 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     6121 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     6181 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     6241 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     6301 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     6361 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     6421 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     6481 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     6541 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     6601 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     6661 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     6721 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     6781 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     6841 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     6901 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     6961 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     7021 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     7081 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     7141 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     7201 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     7261 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     7321 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     7381 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     7441 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     7501 aggggttccg cgcacatttc cccgaaaagt gccacctggg tccttttcat cacgtgctat
     7561 aaaaataatt ataatttaaa ttttttaata taaatatata aattaaaaat agaaagtaaa
     7621 aaaagaaatt aaagaaaaaa tagtttttgt tttccgaaga tgtaaaagac tctaggggga
     7681 tcgccaacaa atactacctt ttatcttgct cttcctgctc tcaggtatta atgccgaatt
     7741 gtttcatctt gtctgtgtag aagaccacac acgaaaatcc tgtgatttta cattttactt
     7801 atcgttaatc gaatgtatat ctatttaatc tgcttttctt gtctaataaa tatatatgta
     7861 aagtacgctt tttgttgaaa ttttttaaac ctttgtttat ttttttttct tcattccgta
     7921 actcttctac cttctttatt tactttctaa aatccaaata caaaacataa aaataaataa
     7981 acacagagta aattcccaaa ttattccatc attaaaagat acgaggcgcg tgtaagttac
     8041 aggcaagcga tccgtctaag aaaccattat tatcatgaca ttaacctata aaaataggcg
     8101 tatcacgagg ccctttcgtc tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat
     8161 gcagctcccg gagacggtca cagcttgtct gtaagcggat gccgggagca gacaagcccg
     8221 tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg cggcatcaga
     8281 gcagattgta ctgagagtgc accataccac cttttcaat
//
