LOCUS       sequence-421719-        7579 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes the S. cerevisiae SUC2 (Invertase) protein (Part3b') with 
            the 111AA SCW4 TFP (Part3a') assembled with pTEF1 promoter, tTDH1 
            terminator, LEU2 selectable marker and CEN/ARS yeast origin..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7579)
  AUTHORS   O'Riordan NM, Juric V, O'Neill SK, Roche AP, Young PW
  TITLE     A Yeast Modular Cloning (MoClo) Toolkit Expansion for Optimization 
            of Heterologous Protein Secretion and Surface Display in 
            Saccharomyces cerevisiae.
  JOURNAL   ACS Synth Biol. 2024 Mar 14. doi: 10.1021/acssynbio.3c00743.
  PUBMED    38483353
REFERENCE   2  (bases 1 to 7579)
  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. 2024 Mar 14. doi: 10.1021/acssynbio.3c00743."
COMMENT     Sequence Label: pYSD113
FEATURES             Location/Qualifiers
     source          1..7579
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        472..885
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             2774..2845
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags"
                     /label=3xFLAG
                     /translation="DYKDDDDKDYKDDDDKDYKDDDDK"
     CDS             2849..2866
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        3521..3928
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             3929..5023
                     /codon_start=1
                     /gene="S. cerevisiae LEU2"
                     /product="3-isopropylmalate dehydrogenase, required for 
                     leucine biosynthesis"
                     /label=LEU2
                     /note="yeast auxotrophic marker"
                     /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL
                     IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA
                     NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT
                     VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ
                     LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF
                     GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG
                     DLGGSNSTTEVGDAVAEEVKKILA"
     misc_feature    5202..5705
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     CDS             complement(join(5816..6607,6608..6676))
                     /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     6439..6458
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6677..6781)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6884..7472
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7373..7392
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ccctgaattc gcatctagaa cggggtcatc acggctcatc atgcgccaaa caaatgtgtg
       61 caatacacgc tcggatgact gcatgatgac cgcactgact ggggacagca gatccaccta
      121 agcctgtgag agaagcagac acccgacaga tcaaggcagt tacgtctcac caaaccagat
      181 gtcaacacag ctacaacgcc ttgccaacag ggagttcttc agagacatgg aggctcaaaa
      241 cgaaattatt gacagcctag acatcaatag tcatacaaca gaaagcgacc acccaacttt
      301 ggctgataat agcgtataaa caatgcatac tttgtacgtt caaaatacaa tgcagtagat
      361 atatttatgc atattacata taatacatat cacataggaa gcaacaggcg cgttggactt
      421 ttaattttcg aggaccgcga atccttacat cacacccaat cccccacaag tgatccccca
      481 cacaccatag cttcaaaatg tttctactcc ttttttactc ttccagattt tctcggactc
      541 cgcgcatcgc cgtaccactt caaaacaccc aagcacagca tactaaattt cccctctttc
      601 ttcctctagg gtgtcgttaa ttacccgtac taaaggtttg gaaaagaaaa aagacaccgc
      661 ctcgtttctt tttcttcgtc gaaaaaggca ataaaaattt ttatcacgtt tctttttctt
      721 gaaaattttt ttttttgatt tttttctctt tcgatgacct cccattgata tttaagttaa
      781 taaacggtca tcaatttctc aagtttcagt ttcatttttc ttgttctatt acaacttttt
      841 ttacttcttg ctcattagaa agaaagcata gcaatctaat ctaagtttta attacaaaag
      901 atctatgcgt ctatctaacc taattgcttc tgcctctctt ttatctgctg ctactcttgc
      961 tgctcccgct aaccacgaac acaaggacaa gcgtgctgtg gtcactacca ctgttcaaaa
     1021 acaaaccact atcattgtta atggtgccgc ttcaactcca gttgctgctt tggaagaaaa
     1081 tgctgttgtc aactccgctc cagctgccgc taccagtaca acatcgtctg ctgcttctgt
     1141 agctaccgct gctgcttcct cttctgagaa caactcacaa gtttctgctg ccgcatctcc
     1201 agcctccagc tctgctgcta catctactca atcttcccta gacaaacgtg cttcaatgac
     1261 aaacgaaact agcgatagac ctttggtcca cttcacaccc aacaagggct ggatgaatga
     1321 cccaaatggg ttgtggtacg atgaaaaaga tgccaaatgg catctgtact ttcaatacaa
     1381 cccaaatgac accgtatggg gtacgccatt gttttggggc catgctactt ccgatgattt
     1441 gactaattgg gaagatcaac ccattgctat cgctcccaag cgtaacgatt caggtgcttt
     1501 ctctggctcc atggtggttg attacaacaa cacgagtggg tttttcaatg atactattga
     1561 tccaagacaa agatgcgttg cgatttggac ttataacact cctgaaagtg aagagcaata
     1621 cattagctat tctcttgatg gtggttacac ttttactgaa taccaaaaga accctgtttt
     1681 agctgccaac tccactcaat tcagagatcc aaaggtgttc tggtatgaac cttctcaaaa
     1741 atggattatg acggctgcca aatcacaaga ctacaaaatt gaaatttact cctctgatga
     1801 cttgaagtcc tggaagctag aatctgcatt tgccaatgaa ggtttcttag gctaccaata
     1861 cgaatgtcca ggtttgattg aagtcccaac tgagcaagat ccttccaaat cttattgggt
     1921 catgtttatt tctatcaacc caggtgcacc tgctggcggt tccttcaacc aatattttgt
     1981 tggatccttc aatggtactc attttgaagc gtttgacaat caatctagag tggtagattt
     2041 tggtaaggac tactatgcct tgcaaacttt cttcaacact gacccaacct acggttcagc
     2101 attaggtatt gcctgggctt caaactggga gtacagtgcc tttgtcccaa ctaacccatg
     2161 gagatcatcc atgtctttgg tccgcaagtt ttctttgaac actgaatatc aagctaatcc
     2221 agagactgaa ttgatcaatt tgaaagccga accaatattg aacattagta atgctggtcc
     2281 ctggtcgcgt tttgctacta acacaactct aactaaggcc aattcttaca atgtcgattt
     2341 gagcaactcg actggtaccc tagagtttga gttggtttac gctgttaaca ccacacaaac
     2401 catatccaaa tccgtctttg ccgacttatc actttggttc aagggtttag aagatcctga
     2461 agaatatttg agaatgggtt ttgaagtcag tgcttcttcc ttctttttgg accgtggtaa
     2521 ctctaaggtc aagtttgtca aggagaaccc atatttcaca aacagaatgt ctgtcaacaa
     2581 ccaaccattc aagtctgaga acgacctaag ttactataaa gtgtacggcc tactggatca
     2641 aaacatcttg gaattgtact tcaacgatgg agatgtggtt tctacaaata cctacttcat
     2701 gaccaccggt aacgctctag gatctgtgaa catgaccact ggtgtcgata atttgttcta
     2761 cattgacgga tccgattaca aggacgatga tgataaggac tacaaagatg atgatgacaa
     2821 agattataag gacgatgatg ataaaggtca tcatcatcat catcactaac tcgagtggca
     2881 taaagcaatc ttgatgagga taatgatttt tttttgaata tacataaata ctaccgtttt
     2941 tctgctagat tttgtgatga cgtaaataag tacatattac tttttaagcc aagacaagat
     3001 taagcattaa ctttaccctt ttctttctaa gtttcaatat tagttatcac tgtttaaaag
     3061 ttatggcgag aacgtcggcg gttaaaatat attaccctga acggctggat gtgagacgat
     3121 aaatcgcagc caagtgagtg aatagatgac gcaccacggt cagacacggc cacatcgtat
     3181 ctcacaggag caagcgcgat aggagcactc acacatagta cggtgatccg ctgactcctt
     3241 tgcccaaata agacgtgagc cactagtgca ctgcagtaca tcgaggagaa cttctagtat
     3301 atctacatac ctaatattat tgccttatta aaaatggaat cccaacaatt acatcaaaat
     3361 ccacattctc ttcaaaatca attgtcctgt acttccttgt tcatgtgtgt tcaaaaacgt
     3421 tatatttata ggataattat actctatttc tcaacaagta attggttgtt tggccgagcg
     3481 gtctaaggcg cctgattcaa gaaatatctt gaccgcagtt aactgtggga atactcaggt
     3541 atcgtaagat gcaagagttc gaatctctta gcaaccatta tttttttcct caacataacg
     3601 agaacacaca ggggcgctat cgcacagaat caaattcgat gactggaaat tttttgttaa
     3661 tttcagaggt cgcctgacgc atataccttt ttcaactgaa aaattgggag aaaaaggaaa
     3721 ggtgagagcg ccggaaccgg cttttcatat agaatagaga agcgttcatg actaaatgct
     3781 tgcatcacaa tacttgaagt tgacaatatt atttaaggac ctattgtttt ttccaatagg
     3841 tggttagcaa tcgtcttact ttctaacttt tcttaccttt tacatttcag caatatatat
     3901 atatatattt caaggatata ccattctaat gtctgcccct aagaagatcg tcgttttgcc
     3961 aggtgaccac gttggtcaag aaatcacagc cgaagccatt aaggttctta aagctatttc
     4021 tgatgttcgt tccaatgtca agttcgattt cgaaaatcat ttaattggtg gtgctgctat
     4081 cgatgctaca ggtgttccac ttccagatga ggcgctggaa gcctccaaga aggctgatgc
     4141 cgttttgtta ggtgctgtgg gtggtcctaa atggggaacc ggtagtgtta gacctgaaca
     4201 aggtttacta aaaatccgta aagaacttca attgtacgcc aacttaagac catgtaactt
     4261 tgcatccgac tctcttttag acttatctcc aatcaagcca caatttgcta aaggtactga
     4321 cttcgttgtt gtcagagaat tagtgggagg tatttacttt ggtaagagaa aggaggacga
     4381 tggtgatggt gtcgcttggg atagtgaaca atacaccgtt ccagaagtgc aaagaatcac
     4441 aagaatggcc gctttcatgg ccctacaaca tgagccacca ttgcctattt ggtccttgga
     4501 taaagctaat gttttggcct cttcaagatt atggagaaaa actgtggagg aaaccatcaa
     4561 gaacgaattt cctacattga aggttcaaca tcaattgatt gattctgccg ccatgatcct
     4621 agttaagaac ccaacccacc taaatggtat tataatcacc agcaacatgt ttggtgatat
     4681 catctccgat gaagcctccg ttatcccagg ttccttgggt ttgttgccat ctgcgtcctt
     4741 ggcctctttg ccagacaaga acaccgcatt tggtttgtac gaaccatgcc acggttctgc
     4801 tccagatttg ccaaagaata aggtcaaccc tatcgccact atcttgtctg ctgcaatgat
     4861 gttgaaattg tcattgaact tgcctgaaga aggtaaggcc attgaagatg cagttaaaaa
     4921 ggttttggat gcaggtatca gaactggtga tttaggtggt tccaacagta ccaccgaagt
     4981 cggtgatgct gtcgccgaag aagttaagaa aatccttgct taacctggag gacccttctc
     5041 tttagactat tctactctta tgcacgtaaa aaattctagg aaatatgtat taactaggag
     5101 taaaataacc ggctagtggc attcatatag ccgtctgttt acatctacat cacacatttc
     5161 gagtgtatat ctcgcaacgt tggcgttaaa taggcaggag tatcacgtgc tataaaaata
     5221 attataattt aaatttttta atataaatat ataaattaaa aatagaaagt aaaaaaagaa
     5281 attaaagaaa aaatagtttt tgttttccga agatgtaaaa gactctaggg ggatcgccaa
     5341 caaatactac cttttatctt gctcttcctg ctctcaggta ttaatgccga attgtttcat
     5401 cttgtctgtg tagaaaacca cacacgaaaa tcctgtgatt ttacatttta cttatcgtta
     5461 atcgaatgta tatctattta atctgctttt cttgtctaat aaatatatat gtaaagtacg
     5521 ctttttgttg aaatttttta aacctttgtt tatttttttt tcttcattcc gtaactcttc
     5581 taccttcttt atttactttc taaaatccaa atacaaaaca taaaaataaa taaacacaga
     5641 gtaaattccc aaattattcc atcattaaaa gatacgaggc gcgtgtaagt tacaggcaag
     5701 cgatccgtcc cgagcggccg cgattatcaa aaaggatctt cacctagatc cttttaaatt
     5761 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     5821 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5881 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5941 ctgcaatgat accgcgggac ccacgctcac cggctccaga tttatcagca ataaaccagc
     6001 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     6061 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     6121 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     6181 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     6241 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     6301 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     6361 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     6421 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     6481 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     6541 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     6601 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     6661 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     6721 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     6781 gcacatttcc ccgaaaagtg ccacctgtca tgaccaaaat cccttaacgt gagttttcgt
     6841 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     6901 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     6961 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     7021 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     7081 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     7141 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     7201 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     7261 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7321 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7381 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7441 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7501 tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg
     7561 tggataaccg tgcggccgc
//
