LOCUS       sequence-245797-        8396 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Integrate pTRX2_5xUAS-CDC14-tADH1 into yeast genome EasyClone site 
            X-4 with SpHIS5 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8396)
  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 8396)
  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: pCfB4613
FEATURES             Location/Qualifiers
     source          1..8396
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..344)
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     protein_bind    complement(399..432)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     terminator      complement(480..669)
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     terminator      3262..3427
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(3317..3341)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     protein_bind    complement(4028..4061)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(4268..4287)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     complement(4293..4312)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        4354..4445
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4446..4514,4515..5306)
                     /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     complement(4664..4683)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5428..6016
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5917..5936
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6170..6187
                     /label=L4440
                     /note="L4440 vector, forward primer"
     primer_bind     6286..6302
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6772..6805
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     7344..7366
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     7359..7375
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     7385..7404
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        7385..7403
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    complement(7447..7480)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            complement(join(7540..8396,1..344))
                     /label=HIS3MX6
                     /note="yeast selectable marker encoding the S. pombe his5 
                     gene, which corresponds to S. cerevisiae HIS3"
     terminator      complement(7540..7737)
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     CDS             complement(7743..8396)
                     /codon_start=1
                     /gene="Schizosaccharomyces pombe his5"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=S. pombe his5
                     /note="yeast auxotrophic marker; corresponds to S. 
                     cerevisiae HIS3"
                     /translation="MGRRAFVERNTNETKISVAIALDKAPLPEESNFIDELITSKHANQ
                     KGEQVIQVDTGIGFLDHMYHALAKHAGWSLRLYSRGDLIIDDHHTAEDTAIALGIAFKQ
                     AMGNFAGVKRFGHAYCPLDEALSRSVVDLSGRPYAVIDLGLKREKVGELSCEMIPHLLY
                     SFSVAAGITLHVTCLYGSNDHHRAESAFKSLAVAMRAATSLTGSSEVPSTKGVL"
ORIGIN
        1 ggttgtttat gttcggatgt gatgtgagaa ctgtatccta gcaagatttt aaaaggaagt
       61 atatgaaaga agaacctcag tggcaaatcc taacctttta tatttctcta caggggcgcg
      121 gcgtggggac aattcaacgc gtctgtgagg ggagcgtttc cctgctcgca ggtctgcagc
      181 gaggagccgt aatttttgct tcgcgccgtg cggccatcaa aatgtatgga tgcaaatgat
      241 tatacatggg gatgtatggg ctaaatgtac gggcgacagt cacatcatgc ccctgagctg
      301 cgcacgtcaa gactgtcaag gagggtattc tgggcctcca tgtcgctggc cgggtgaccc
      361 ggcggggacg aggcaagcta aacagatctc tagacctaat aacttcgtat agcatacatt
      421 atacgaagtt atattaaggg ttgtcgacct gcagcgtacg aagcttcagc tgacgcgatc
      481 ttcgagcgtc ccaaaacctt ctcaagcaag gttttcagta taatgttaca tgcgtacacg
      541 cgtctgtaca gaaaaaaaag aaaaatttga aatataaata acgttcttaa tactaacata
      601 actataaaaa aataaatagg gacctagact tcaggttgtc taactccttc cttttcggtt
      661 agagcggatg aatgcacgcg atcgtgcgat cttgagtgcc agtgaatatt aagtaatcat
      721 caaagtatat gtgtaattgt ttatactctt agtaaaggat gctccctaca aggtggctct
      781 tttcttacta agcgcgttca gtttccagcc agccgaaaga gggatatcag cttgagtgcc
      841 agtgaatatt aagtaatcat caaagtatat gtgtaattgt ttatactctt agtaaaggat
      901 gctccctaca aggtggctct tttcttacta agcgcgttca gtttccagcc agccgaaaga
      961 gggatatcag acctgcactc ttgagtgcca gtgaatatta agtaatcatc aaagtatatg
     1021 tgtaattgtt tatactctta gtaaaggatg ctccctacaa ggtggctctt ttcttactaa
     1081 gcgcgttcag tttccagcca gccgaaagag ggatatcagc ttgagtgcca gtgaatatta
     1141 agtaatcatc aaagtatatg tgtaattgtt tatactctta gtaaaggatg ctccctacaa
     1201 ggtggctctt ttcttactaa gcgcgttcag tttccagcca gccgaaagag ggatatcaga
     1261 tctgtcatct tgagtgccag tgaatattaa gtaatcatca aagtatatgt gtaattgttt
     1321 atactcttag taaaggatgc tccctacaag gtggctcttt tcttactaag cgcgttcagt
     1381 ttccagccag ccgaaagagg gatatcagta tataagaaag ccattcgggg gatgaaaagc
     1441 tgacaagaga ataacgagga ccagttttta tttgttgtct agcaagaatt atacacgcac
     1501 acatacacga gagtctacga tatctttaaa taacacatca ataagtgcag gtaaaacaat
     1561 gcgtaggagt gtatacctcg acaacacgat cgagttcctg cgaggcagag tatatctcgg
     1621 tgcatatgat tacacgccgg aagatactga tgagctggtg ttttttaccg tggaggacgc
     1681 aatcttttac aacagtttcc atttggattt cggtcccatg aacattggac atctatatcg
     1741 ttttgccgtt attttccatg aaatcttgaa tgacccagaa aatgccaata aagccgttgt
     1801 tttctattca tctgcatcga ccagacaacg tgctaacgcc gcatgtatgc tttgctgtta
     1861 tatgatcctc gttcaggcat ggactccgca tcaagtgctt cagccgttgg cgcaagtaga
     1921 ccctcccttc atgccattta gagatgccgg ttattctaat gcggatttcg aaatcacaat
     1981 tcaagatgtg gtttatggcg tttggagggc taaggaaaaa ggccttattg atttgcattc
     2041 cttcaatttg gagagttacg agaaatacga acacgtggag tttggagact tcaatgtact
     2101 caccccagat tttattgcat ttgcgtcacc acaagaggac catccaaagg gctatctcgc
     2161 taccaaatct tcacatttga accagccgtt taagtcggtt ctgaattttt tcgccaacaa
     2221 taacgtccaa ttagtggtaa ggttgaattc tcatttatat aataaaaagc attttgaaga
     2281 cattggcatt caacatttgg atctaatttt cgaagatggt acatgtcctg atctttccat
     2341 tgtaaaaaac tttgttggtg ctgctgaaac gataattaaa aggggaggga aaattgctgt
     2401 gcactgtaag gctgggctag gtagaactgg ttgtttaatt ggtgctcact taatatacac
     2461 gtatggattt accgctaacg agtgtattgg cttcttaagg tttataaggc cgggaatggt
     2521 cgttggccct caacagcatt ggttatattt acaccagaat gattttagag aatggaagta
     2581 cacaaccaga atttctttga agcctagtga ggccattggc ggcttatatc ccttaataag
     2641 tttagaagaa taccgcttac aaaaaaagaa acttaaagac gacaagaggg ttgcgcaaaa
     2701 taacattgaa ggtgaattaa gagatttaac catgacgccg ccatccaacg gtcatggggc
     2761 gcttagtgcc agaaattcga gccaaccttc tacggcgaat aatggaagta attcattcaa
     2821 gagctcagcc gtcccacaga cttcgcctgg tcagccgaga aagggccaaa acggttctaa
     2881 tactatagaa gatatcaaca ataatcgtaa tccaacttcc catgcgaata ggaaggtagt
     2941 aatagaaagc aacaactcgg atgacgaatc catgcaggat accaacggca caagtaatca
     3001 ttatcccaaa gtttcgcgca agaaaaatga tatctcctct gcttcatcgt cccgaatgga
     3061 agataacgaa cccagcgcaa cgaacatcaa taatgccgca gatgatacca tactaagaca
     3121 actattgccc aagaatagga gagtaacgtc agggagaagg actacaagcg ccgccggtgg
     3181 tataagaaaa ataagtggct ccatcaagaa ataaatcgca cgcattccgt tggtagatac
     3241 gttgttgaca cttctaaata agcgaatttc ttatgattta tgatttttat tattaaataa
     3301 gttataaaaa aaataagtgt atacaaattt taaagtgact cttaggtttt aaaacgaaaa
     3361 ttcttattct tgagtaactc tttcctgtag gtcaggttgc tttctcaggt atagcatgag
     3421 gtcgctcatc gcacgcattc catgcgagct cgctgaggac ttaatctgct cttgaatggc
     3481 gacagcctat tgccccagtg ttccctcaac aaccttggta gttggagcgc aattagcgta
     3541 tcctgtacca tactaattct cttctgccca acgacggctg ccattagtca gcatggcgcg
     3601 cacgtgacta caactgtggc tggaaacctt ttcgtcctcc ccggtttttc agtgagccga
     3661 ctctactaca atgctttttc atttttcact cagaaaaacc tgcaatttgc caaattggcc
     3721 atgctctgtg cctcccttga caaaggacat cttccctgtt tataaacggc ggcttaccaa
     3781 aagttgaagc ttgttcttgc ctcttatgag tggagcaatc gattatattg aatcgttgtg
     3841 ctggagtagt tggatctttc cacgtggtct cgagtcactt gtagaagctg aaaattgagc
     3901 agatttagta tagggctaca ttgtagggtg gtttagagta tcgaaaatat acatatagaa
     3961 gaataaaatg ggactcttag ctttgggatt aaaccctcag cgcggccgca tttaaatccc
     4021 cacttcagaa gttcctatac tttctagaga ataggaactt cactatagag tcgaataagg
     4081 gcgacacccc ataattagcc cgggcgaaag gcccagtctt tcgactgagc ctttcgtttt
     4141 atttgatgcc tggcagttcc ctactctcgc atggggagtc cccacactac catcggcgct
     4201 acggcgtttc acttctgagt tcggcatggg gtcaggtggg accaccgcgc tactgccgcc
     4261 aggcaaacaa ggggtgttat gagccatatt caggtataaa tgggctcgcg ataatgttca
     4321 gaattggtta attggttgta acactgaccc ctatttgttt atttttctaa atacattcaa
     4381 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4441 agaatatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     4501 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     4561 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     4621 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     4681 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     4741 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     4801 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     4861 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     4921 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     4981 cgatgcctgt agcgatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     5041 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     5101 tgcgctcggc ccttccggct ggctggttta ttgctgataa atccggagcc ggtgagcgtg
     5161 gttctcgcgg tatcatcgca gcgctggggc cagatggtaa gccctcccgt atcgtagtta
     5221 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     5281 gtgcctcact gattaagcat tggtaagcag agcattacgc tgacttgacg ggacggcgca
     5341 agctcatgac caaaatccct taacgtgagt tacgcgcgcg tcgttccact gagcgtcaga
     5401 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     5461 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     5521 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     5581 agtgtagccg tagttagccc accacttcaa gaactctgta gcaccgccta catacctcgc
     5641 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     5701 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     5761 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     5821 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     5881 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     5941 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     6001 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     6061 gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac
     6121 cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt
     6181 gagcgaggaa gcggaaggcg agagtaggga actgccaggc atcaaactaa gcagaaggcc
     6241 cctgacggat ggcctttttg cgtttctaca aactctttct gtgttgtaaa acgacggcca
     6301 gtcttaagct cgggccccct gggcggttct gataacgagt aatcgttaat ccgcaaataa
     6361 cgtaaaaacc cgcttcggcg ggttttttta tggggggagt ttagggaaag agcatttgtc
     6421 agaatattta agggcgcctg tcactttgct tgatatatga gaattattta accttataaa
     6481 tgagaaaaaa gcaacgcact ttaaataaga tacgttgctt tttcgattga tgaacaccta
     6541 taattaaact attcatctat tatttatgat tttttgtata tacaatattt ctagtttgtt
     6601 aaagagaatt aagaaaataa atctcgaaaa taataaaggg aaaatcagtt tttgatatca
     6661 aaattataca tgtcaacgat aatacaaaat ataatacaaa ctataagatg ttatcagtat
     6721 ttattatgca tttagaataa attttgtgtc gcccttattc gactcactat agaagttcct
     6781 attctctaga aagtatagga acttcacttc attttattta aatttgcggc cgcgctgagg
     6841 tcttaatctg gtgaggattt acggtatgat catgctggta agcttcgtta ctcatgcttt
     6901 ggttggcttc ttgaaagtgt ctagttcact ttatgcccgg ttataaacat aaactttcac
     6961 agggttctct ttgatcacct ctttgtttaa tcttatgaat ccaagggatt ttatcggtca
     7021 aaggggaaat caatgcgaaa gacagtaatg atgagagaaa aactctccgt aaccaccaag
     7081 tttggttcag cgcgacgaga tttttatcga ttatcgagaa aaatacctgt atatctacat
     7141 ttctatgtca gtgatatata cttcttagat aagttatgcc accagtgcat acgcttacgc
     7201 acacacacgt attcttgtgc acacgcctgt tacttcttgc agacatcaga catactattg
     7261 taattcaaaa aaaaaaagcg aatcttccca tgcctgttat taatgcctca gcactagtcc
     7321 tgcaggggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgagcg
     7381 cgcgtaatac gactcactat agggcgaatt gggtgcatag gccactagtg gatctgatat
     7441 cacctaataa cttcgtatag catacattat acgaagttat attaagggtt ctcgagagct
     7501 cgtttaaact ggatggcggc gttagtatcg aatcgacagc agtatagcga ccagcattca
     7561 catacgattg acgcatgata ttactttctg cgcacttaac ttcgcatctg ggcagatgat
     7621 gtcgaggcga aaaaaaatat aaatcacgct aacatttgat taaaatagaa caactacaat
     7681 ataaaaaaac tatacaaatg acaagttctt gaaaacaaga atctttttat tgtcagtact
     7741 ctttacaaca ctcccttcgt gcttgggact tcagaacttc cagtaagact agtagccgcg
     7801 cgcatggcaa cagccagaga tttaaaagcg ctttcagcac gatgatggtc attactacca
     7861 tataagcagg taacatgcaa agtaattcca gctgctaccg aaaaggaata tagtaagtga
     7921 gggatcattt cacaggacaa ttccccaacc ttttcacgct ttaatcccaa atcgataaca
     7981 gcatagggcc gtcccgacaa gtcaactacg cttctagaaa gggcttcgtc aagtggacaa
     8041 taagcatgtc caaatctttt aacgccggca aagttaccca tagcctgctt gaatgcaata
     8101 ccaagtgcaa tagcagtatc ttctgcagtg tgatgatcat cgatgattaa atcacctctt
     8161 gagtaaagtc gtaagctcca gcctgcatgt ttagccagtg catgatacat gtgatccagg
     8221 aatccaattc ccgtgtctac ttggattact tgttctccct tttggtttgc atgcttggaa
     8281 gttataagtt catcaataaa attcgactct tcaggtaagg gagctttgtc caaagcgatg
     8341 gcaacgctga ttttcgtttc gttcgtattt ctttctacaa aagccctcct acccat
//
