LOCUS       sequence-384740-        6554 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid for genome integration in X2 site expressing PyruEth 
            (Pyruvate and Ethanol consumption sensors).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6554)
  AUTHORS   Torello Pianale L, Olsson L
  TITLE     ScEnSor Kit for Saccharomyces cerevisiae Engineering and 
            Biosensor-Driven Investigation of the Intracellular Environment.
  JOURNAL   ACS Synth Biol. 2023 Aug 8. doi: 10.1021/acssynbio.3c00124.
  PUBMED    37552581
REFERENCE   2  (bases 1 to 6554)
  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: "ACS Synth 
            Biol. 2023 Aug 8. doi: 10.1021/acssynbio.3c00124."
COMMENT     Sequence Label: LT2_32_PyruEth_X2site
FEATURES             Location/Qualifiers
     source          1..6554
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(1397..1413)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2859..2875)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3146..3543
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             complement(3442..3453)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             3557..4267
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVDGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(3702..3720)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     3970..3989
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     terminator      4281..4468
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     complement(4336..4360)
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(4717..5532)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(4717..5532)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5440..5459
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5524..5543
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      5757..6345
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6246..6265
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tgccctgaat tcgcatctag actgaactgg ccgataattg cagacgaacg gtagcttcca
       61 gtttttggtg gttctggctt ccttcccgat tccgcccgct aaacgcataa ttttgttgcc
      121 tggtggcatt tgcaaaatgc ataacctatg catttaaaag attatgtatg ctcttctgac
      181 ttttcgtgtg atgaggctcg tggaaaaaat gaataattta tgaatttgag aacaattttg
      241 tgttgttacg gtattttact atggaataat caatcaattg aggattttat gcaaatatcg
      301 tttgaatatt tttccgaccc tttgagtact tttcttcata attgcataat attgtccgct
      361 gcccgttttt ctgttagacg gtgtcttgat ctacttgcta tcgttcaaca ccaccttatt
      421 ttctaactat ttttttttta gctcatttga atcagcttat ggtgatggca catttttgca
      481 taaacctagc tgtcctcgtt gaacatagga aaaaaaaata tataaacaag gctctttcac
      541 tctccttgga atcagatttg ggtttgttcc ctttattttc atatttcttg tcatattctt
      601 ttctcaatta ttatcttcta ctcataacct cacgcaaaat aacacagtca aatcaatcaa
      661 atatggtatc caagggcgaa gaattgttca ccggtgtggt cccaatcttg gtcgagttgg
      721 acggtgacgt caacggtcac aagttctctg tttccggtga aggtgaagga gatgccacct
      781 acggtaaatt gaccttgaag ttgttgtgca ccaccggcaa gctgccagtt ccatggccaa
      841 cattggtcac cactttaggt tatggtttac aatgttttgc tagatatcca gatcatatga
      901 aacaacatga tttcttcaaa tctgctatgc cagaaggtta tgttcaagaa agaactatat
      961 ttttcaaaga tgatggtaat tataaaacta gagctgaagt taaatttgaa ggagatactt
     1021 tagttaatag aattgaattg aaaggtattg attttaaaga agatggtaat attttaggtc
     1081 ataaattaga atataattat aattctcata atgtttatat tacagctgat aaacaaaaga
     1141 atggtattaa ggctaatttt aaaattagac ataatattga agatggtggt gttcaattag
     1201 ctgatcatta tcaacaaaat actcctattg gtgacggtcc agttttatta cctgataatc
     1261 attatttgtc atatcaatct aaattgtcaa aagatccaaa tgaaaaaaga gatcatatgg
     1321 ttttgttaga atttttgact gcagctggta ttactttagg tatggatgaa ttgtataaag
     1381 gatcctaact cgagatcgat accgtcgacc tcgagtcatg taattagtta tgtcacgctt
     1441 acattcacgc cctcctccca catccgctct aaccgaaaag gaaggagtta gacaacctga
     1501 agtctaggtc cctatttatt ttttttaata gttatgttag tattaagaac gttatttata
     1561 tttcaaattt ttcttttttt tctgtacaaa cgcgtgtacg catgtaacat tatactgaaa
     1621 accttgcttg agaaggtttt gggacgctcg gctgccaaac cagatgtcaa cacagctaca
     1681 acgtcctgcc tttctaatca ccattctaat gttttaatta agggattttg tcttcattaa
     1741 cggctttcgc tcataaaaat gttatgacgt tttgcccgca ggcgggaaac catccacttc
     1801 acgagactga tctcctctgc cggaacaccg ggcatctcca acttataagt tggagaaata
     1861 agagaatttc agattgagag aatgaaaaaa aaaaaaaaaa aggcagagga gagcataaaa
     1921 atggggttca ctttttggta aagctatagc atgcctatca catataaata gagtgccagt
     1981 agcgactttt ttcacactcg aaatactctt actactgctc tcttgttgtt tttatcactt
     2041 cttgtttctt cttggtaaat agaatatcaa gctacaaaaa gcatacaatc aactatcaac
     2101 tattaactat atcgtaatac acatatggtt tctaaaggtg aagaattatt cactggtgtt
     2161 gtcccaattt tggttgaatt agatggtgat gttaatggtc acaaattttc tgtctccggt
     2221 gaaggtgaag gtgatgctac ttacggtaaa ttgaccttaa aatttatttg tactactggt
     2281 aaattgccag ttccatggcc aaccttagtc actactttat cttggggtgt tcaatgtttt
     2341 gcaagatacc cagatcatat gaaacaacat gactttttca agtctgccat gccagaaggt
     2401 tatgttcaag aaagaactat ttttttcaaa gatgacggta actacaagac cagagctgaa
     2461 gtcaagtttg aaggtgatac cttagttaat agaatcgaat taaaaggtat tgattttaaa
     2521 gaagatggta acattttagg tcacaaattg gaatacaatt atttctctga caatgtttac
     2581 atcactgctg acaaacaaaa gaatggtatc aaagctaact tcaaaattag acacaacatt
     2641 gaagatggtg gtgttcaatt agctgaccat tatcaacaaa atactccaat tggtgatggt
     2701 ccagtcttgt taccagacaa ccattactta tccactcaat ctaagttatc caaagatcca
     2761 aacgaaaaga gggaccacat ggtcttgtta gaatttgtta ctgctgctgg tattaccttg
     2821 ggtatggatg aattgtacaa aggatcctaa ctcgagatcg ataccgtcga cctcgagtca
     2881 tgtaattagt tatgtcacgc ttacattcac gccctcctcc cacatccgct ctaaccgaaa
     2941 aggaaggagt tagacaacct gaagtctagg tccctattta ttttttttaa tagttatgtt
     3001 agtattaaga acgttattta tatttcaaat ttttcttttt tttctgtaca aacgcgtgta
     3061 cgcatgtaac attatactga aaaccttgct tgagaaggtt ttgggacgct cggctggatg
     3121 cacacactgg cttaagatga caacgatagc ttcgaaacgt ttctactcct cttttactct
     3181 tccagatctt ctcggactcc gcgcatcgcc gtaccacttc aaaacaccca agcacagcat
     3241 actaaatttc ccctctttct tcctctaggg tgtcgttaat tacccgtact aaaggtttgg
     3301 aaaagaaaaa agagaccgcc tcgtttcttt ttcttcgtcg aaaaaggcaa taaaaatttt
     3361 tatcacgttt ctttttcttg aaaattcttt ttttgatttt tttctctttc gatgacctcc
     3421 catcgatatt taagttaata aacggccttc aatttcccaa gtttcagtcc cacttttctt
     3481 gctctactac aacttttttt acctcttgct cattagaaag aaagcatagc aatctaatct
     3541 aagttttcta gaaaatatgg tgagcaaggg cgaggaggat aacatggcca tcatcaagga
     3601 gttcatgcgc ttcaaggtgc acatggaggg ctccgtggac ggccacgagt tcgagatcga
     3661 gggcgagggc gagggccgcc cctacgaggg cacccagacc gccaagctga aggtgaccaa
     3721 gggtggcccc ctgcccttcg cctgggacat cctgtcccct cagttcatgt acggctccaa
     3781 ggcctacgtg aagcaccccg ccgacatccc cgactacttg aagctgtcct tccccgaggg
     3841 cttcaagtgg gagcgcgtga tgaacttcga ggacggcggc gtggtgaccg tgacccagga
     3901 ctcctccctg caggacggcg agttcatcta caaggtgaag ctgcgcggca ccaacttccc
     3961 ctccgacggc cccgtaatgc agaagaagac tatgggctgg gaggcctcct ccgagcggat
     4021 gtaccccgag gacggcgccc tgaagggcga gatcaagcag aggctgaagc tgaaggacgg
     4081 cggccactac gacgctgagg tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc
     4141 cggcgcctac aacgtcaaca tcaagttgga catcacctcc cacaacgagg actacaccat
     4201 cgtggaacag tacgaacgcg ccgagggccg ccactccacc ggcggcatgg acgagctgta
     4261 caagtaaatc ctaactcgag gcgaatttct tatgatttat gatttttatt attaaataag
     4321 ttataaaaaa aataagtgta tacaaatttt aaagtgactc ttaggtttta aaacgaaaat
     4381 tcttattctt gagtaactct ttcctgtagg tcaggttgct ttctcaggta tagcatgagg
     4441 tcgctcttat tgaccacacc tctaccggca tgccgagcaa atgcctgcaa atcgctcccc
     4501 atttcgctga gcagttacag agatgttacg aaccactagt gcactgcagt acataacccc
     4561 taggtcttcc tgagagtgac gaaggctaag gtcacttctt cgtttccttt attggggttt
     4621 ccgtgtagcc ttcccctgaa tagtgtggga cgttttatga gaagccgtaa gaaatagccg
     4681 agcggccgcg tgttacaacc aattaaccaa ttctgattag aaaaactcat cgagcatcaa
     4741 atgaaactgc aatttattca tatcaggatt atcaatacca tatttttgaa aaagccgttt
     4801 ctgtaatgaa ggagaaaact caccgaggca gttccatagg atggcaagat cctggtatcg
     4861 gtctgcgatt ccgactcgtc caacatcaat acaacctatt aatttcccct cgtcaaaaat
     4921 aaggttatca agtgagaaat caccatgagt gacgactgaa tccggtgaga atggcaaaag
     4981 cttatgcatt tctttccaga cttgttcaac aggccagcca ttacgctcgt catcaaaatc
     5041 actcgcatca accaaaccgt tattcattcg tgattgcgcc tgagcgaggc gaaatacgcg
     5101 atcgctgtta aaaggacaat tacaaacagg aatcgaatgc aaccggcgca ggaacactgc
     5161 cagcgcatca acaatatttt cacctgaatc aggatattct tctaatacct ggaatgctgt
     5221 tttcccgggg atcgcagtgg tgagtaacca tgcatcatca ggagtacgga taaaatgctt
     5281 gatggtcgga agaggcataa attccgtcag ccagtttagt ctgaccatct catctgtaac
     5341 atcattggca acgctacctt tgccatgttt cagaaacaac tctggcgcat cgggcttccc
     5401 atacaatcga tagattgtcg cacctgattg cccgacatta tcgcgagccc atttataccc
     5461 atataaatca gcatccatgt tggaatttaa tcgcggcctg gagcaagacg tttcccgttg
     5521 aatatggctc ataacacccc ttgtattact gtttatgtaa gcagacagtt ttattgttca
     5581 tgatgatata tttttatctt gtgcaatgta acatcagaga ttttgagaca caacgtggct
     5641 ttgttgaata aatcgaactt ttgctgagtt gaaggatcag tcatgaccaa aatcccttaa
     5701 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     5761 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     5821 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     5881 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     5941 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     6001 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     6061 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     6121 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     6181 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     6241 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     6301 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     6361 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
     6421 tcccctgatt ctgtggataa ccgtgcggcc gccaatgtca aaagatcctc tcataccata
     6481 ttaagtaaat tgcctccatt tctttttcct cgggcagaga aactcgcagg caacttgctc
     6541 tcgaagtggt cacg
//
