LOCUS       sequence-431851-        5474 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast integrative vector for constitutive expression of LDH at the 
            TRP1 locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5474)
  AUTHORS   Choi B, Tafur Rangel A, Kerkhoven EJ, Nygard Y
  TITLE     Engineering of Saccharomyces cerevisiae for enhanced metabolic 
            robustness and L-lactic acid production from lignocellulosic 
            biomass.
  JOURNAL   Metab Eng. 2024 Jul;84:23-33. doi: 10.1016/j.ymben.2024.05.003. Epub
            2024 May 23.
  PUBMED    38788894
REFERENCE   2  (bases 1 to 5474)
  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: "Metab 
            Eng."; date: "2024-07"; volume: "84:23-33. doi"; pages: " 
            10.1016/j.ymben.2024.05.003. Epub 2024 May 23"
COMMENT     Sequence Label: YIplac204-TDH3p-LDH-CYC1t
FEATURES             Location/Qualifiers
     source          1..5474
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    106..127
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(142..147,148..165,166..172)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    180..196
                     /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)."
     primer_bind     185..207
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     204..220
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     204..220
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        252..895
                     /gene="S. cerevisiae TDH3"
                     /label=GAP promoter
                     /note="promoter for glyceraldehyde-3-phosphate 
                     dehydrogenase; also known as the TDH3 promoter"
     primer_bind     788..811
                     /label=GPDpro-F
                     /note="S. cerevisiae GPD promoter, forward primer"
     terminator      1965..2212
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(1981..1999)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2219..2236)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2219..2235)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2228..2250)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(2444..2463)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2563..2585
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(2704..3378)
                     /codon_start=1
                     /gene="S. cerevisiae TRP1"
                     /product="phosphoribosylanthranilate isomerase, required 
                     for tryptophan biosynthesis"
                     /label=TRP1
                     /note="yeast auxotrophic marker"
                     /translation="MSVINFTGSSGPLVKVCGLQSTEAAECALDSDADLLGIICVPNRK
                     RTIDPVIARKISSLVKAYKNSSGTPKYLVGVFRNQPKEDVLALVNDYGIDIVQLHGDES
                     WQEYQEFLGLPVIKRLVFPKDCNILLSAASQKPHSFIPLFDSEAGGTGELLDWNSISDW
                     VGRQESPESLHFMLAGGLTPENVGDALRLNGVIGVDVSGGVETNGVKDSNKIANFVKNA
                     KK"
     promoter        complement(3379..3480)
                     /gene="S. cerevisiae TRP1"
                     /label=TRP1 promoter
     primer_bind     complement(3482..3500)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3568..3672
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3673..3741,3742..4533)
                     /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(3891..3910)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4704..5292
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5193..5212
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5446..5463
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 gcgcccaata cgcaaaccgc ctctccccgc gcgttggccg attcattaat gcagctggca
       61 cgacaggttt cccgactgga aagcgggcag tgagcgcaac gcaattaatg tgagttagct
      121 cactcattag gcaccccagg ctttacactt tatgcttccg gctcgtatgt tgtgtggaat
      181 tgtgagcgga taacaatttc acacaggaaa cagctatgac catgattacg ccaagcttca
      241 gttcgagttt atcattatca atactgccat ttcaaagaat acgtaaataa ttaatagtag
      301 tgattttcct aactttattt agtcaaaaaa ttagcctttt aattctgctg taacccgtac
      361 atgcccaaaa tagggggcgg gttacacaga atatataaca tcgtaggtgt ctgggtgaac
      421 agtttattcc tggcatccac taaatataat ggagcccgct ttttaagctg gcatccagaa
      481 aaaaaaagaa tcccagcacc aaaatattgt tttcttcacc aaccatcagt tcataggtcc
      541 attctcttag cgcaactaca gagaacaggg gcacaaacag gcaaaaaacg ggcacaacct
      601 caatggagtg atgcaacctg cctggagtaa atgatgacac aaggcaattg acccacgcat
      661 gtatctatct cattttctta caccttctat taccttctgc tctctctgat ttggaaaaag
      721 ctgaaaaaaa aggttgaaac cagttccctg aaattattcc cctacttgac taataagtat
      781 ataaagacgg taggtattga ttgtaattct gtaaatctat ttcttaaact tcttaaattc
      841 tacttttata gttagtcttt tttttagttt taaaacacca agaacttagt ttcgaataaa
      901 cacacctgca ggtcgactct agagcatggc gacacttaag gatcagttga tccaaaatct
      961 actgaaagaa gaacacgtac cacagaacaa gataaccatc gtcggggtcg gtgccgtcgg
     1021 aatggcctgc gcaataagta tcttaatgaa agacttagcg gacgaagttg cactggtgga
     1081 tgttatggag gacaaactta agggcgaaat gatggatctg cagcacggat cattgttctt
     1141 acgtacgccc aaaatcgtta gtggtaagga ctataatgta acagctaata gcaggttggt
     1201 aatcattact gccggagcca ggcaacaaga aggcgaatca aggttaaatc tggtacagag
     1261 aaatgtgaac atttttaagt tcataatccc aaatatcgtt aagtatagcc ccaattgcaa
     1321 acttctagta gtgtcaaacc cggtggatat cctaacatac gtggcctgga aaatctcagg
     1381 gttcccgaaa aatcgtgtaa ttggatcagg atgcaacctg gattccgcca gattccgtta
     1441 ccttatgggc gagaggctgg gtgtccatcc tctaagttgc cacggatgga ttttagggga
     1501 gcatggcgac agttcagtac ccgtctggag cggggtcaac gtagctggtg tcagtcttaa
     1561 gaatttgcac ccggagctgg gtactgacgc cgacaaggaa caatggaaag ctgtgcataa
     1621 gcaagtagtt gattccgctt acgaagtgat taagttaaag ggttatacaa gttgggccat
     1681 aggcctatcc gtcgcagatt tagcagagag cattatgaaa aaccttcgtc gtgtccatcc
     1741 gatcagcaca atgattaaag gtttgtatgg gattaaagaa gacgttttct tgagtgttcc
     1801 ttgcatacta gggcaaaatg gtatctcaga tgtcgtcaaa gtgaccctta ctcacgaaga
     1861 agaggcatgt cttaagaagt ctgcagacac cctttgggga attcagaagg aactacagtt
     1921 ctaacccccg ggtaccgagc tcggcccctt ttcctttgtc gatatcatgt aattagttat
     1981 gtcacgctta cattcacgcc ctccccccac atccgctcta accgaaaagg aaggagttag
     2041 acaacctgaa gtctaggtcc ctatttattt ttttatagtt atgttagtat taagaacgtt
     2101 atttatattt caaatttttc ttttttttct gtacaaacgc gtgtacgcat gtaacattat
     2161 actgaaaacc ttgcttgaga aggttttggg acgctcgaag gctttaattt gcgaattcac
     2221 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     2281 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     2341 cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta
     2401 cgcatctgtg cggtatttca caccgcatat ggtgcactct cagtacaatc tgctctgatg
     2461 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     2521 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     2581 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcgatctt ttatgcttgc
     2641 ttttcaaaag gcttgcaggc aagtgcacaa acaatactta aataaatact actcagtaat
     2701 aacctatttc ttagcatttt tgacgaaatt tgctattttg ttagagtctt ttacaccatt
     2761 tgtctccaca cctccgctta catcaacacc aataacgcca tttaatctaa gcgcatcacc
     2821 aacattttct ggcgtcagtc caccagctaa cataaaatgt aagctctcgg ggctctcttg
     2881 ccttccaacc cagtcagaaa tcgagttcca atccaaaagt tcacctgtcc cacctgcttc
     2941 tgaatcaaac aagggaataa acgaatgagg tttctgtgaa gctgcactga gtagtatgtt
     3001 gcagtctttt ggaaatacga gtcttttaat aactggcaaa ccgaggaact cttggtattc
     3061 ttgccacgac tcatctccat gcagttggac gatatcaatg ccgtaatcat tgaccagagc
     3121 caaaacatcc tccttaggtt gattacgaaa cacgccaacc aagtatttcg gagtgcctga
     3181 actattttta tatgctttta caagacttga aattttcctt gcaataaccg ggtcaattgt
     3241 tctctttcta ttgggcacac atataatacc cagcaagtca gcatcggaat ctagtgcaca
     3301 ttctgcggcc tctgtgctct gcaagccgca aactttcacc aatggaccag aactacctgt
     3361 gaaattaata acagacatac tccaagctgc ctttgtgtgc ttaatcacgt atactcacgt
     3421 gctcaatagt caccaatgcc ctccctcttg gccctctcct tttctttttt cgaccgaatt
     3481 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     3541 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     3601 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     3661 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     3721 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     3781 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     3841 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     3901 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     3961 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     4021 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     4081 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     4141 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     4201 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     4261 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     4321 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     4381 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     4441 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     4501 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     4561 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     4621 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     4681 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     4741 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     4801 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     4861 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     4921 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     4981 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     5041 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     5101 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     5161 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     5221 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     5281 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     5341 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
     5401 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
     5461 gaggaagcgg aaga
//
