LOCUS       sequence-421768-        7429 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes the Aga2 signal peptide, mRuby2 fluorescent protein 
            HA-tagged Sed1p yeast surface display anchor protein 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 7429)
  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 7429)
  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: pYSD313
FEATURES             Location/Qualifiers
     source          1..7429
                     /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             962..1669
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="VSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTMR
                     IKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGGV
                     VTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTHM
                     ALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAVA
                     KFAGLGGGMDELYK"
     CDS             1706..1732
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1736..1756
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     promoter        3371..3778
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     CDS             3779..4873
                     /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    5052..5555
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     CDS             complement(join(5666..6457,6458..6526))
                     /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     6289..6308
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6527..6631)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6734..7322
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7223..7242
                     /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 atctatgatg cagttacttc gctgtttttc aatattttct gttattgctt cagttcttgc
      961 tgtgtccaaa ggagaggagt taatcaagga aaacatgaga atgaaagttg tcatggaggg
     1021 ctccgttaat ggtcaccaat tcaagtgtac aggggaaggt gaaggtaatc cttacatggg
     1081 tacacaaact atgagaatta aagtaattga aggcggacca ctaccatttg catttgacat
     1141 tctggcaacg tcattcatgt acggatcacg aactttcatc aagtacccta aaggtatacc
     1201 agactttttc aagcaatctt ttccagaggg ttttacatgg gaaagggtta caagatacga
     1261 agatgggggt gtcgtcacag ttatgcaaga tacttcatta gaagatggct gccttgtcta
     1321 tcatgtgcaa gtaagagggg tgaattttcc ttctaacgga cctgtgatgc agaaaaagac
     1381 caaaggttgg gaaccaaata ctgaaatgat gtacccagct gatggaggtt tgagaggcta
     1441 cacacacatg gcgcttaaag ttgatggtgg aggtcatttg tcttgtagtt ttgttaccac
     1501 ttatcgttct aaaaagactg ttggcaatat caaaatgcca ggaatacatg ctgtagacca
     1561 cagactagaa agactcgaag agagcgataa cgaaatgttc gttgtacaga gagagcatgc
     1621 cgtagccaaa tttgctggct taggcggtgg tatggatgaa ttgtataagg gatccggtgg
     1681 agggggttct ggtggtggcg gatcataccc atacgatgtt ccagattacg ctaaggaaaa
     1741 tttgtatttt caatctcaat tttccaacag tacatctgct tcttccaccg atgtcacttc
     1801 ctcctcttcc atctccactt cctctggctc agtaactatc acatcttctg aagctccaga
     1861 atccgacaac ggtaccagca cagctgcacc aactgaaacc tcaacagagg ctccaaccac
     1921 tgctatccca actaacggta cctctactga agctccaacc actgctatcc caactaacgg
     1981 tacctctact gaagctccaa ctgatactac tactgaagct ccaaccaccg ctcttccaac
     2041 taacggtact tctactgaag ctccaactga tactactact gaagctccaa ccaccggtct
     2101 tccaaccaac ggtaccactt cagctttccc accaactaca tctttgccac caagcaacac
     2161 taccaccact cctccttaca acccatctac tgactacacc actgactaca ctgtagtcac
     2221 tgaatatact acttactgtc cagaaccaac cactttcacc acaaacggta agacttacac
     2281 cgtcactgaa ccaaccacat tgactatcac tgactgtcca tgcaccattg aaaagccaac
     2341 aaccacatca accaccgaat acactgtagt cactgagtac actacttact gtccagaacc
     2401 aaccactttc accacaaacg gtaagactta caccgtcact gaaccaacca ctttgactat
     2461 cactgactgt ccatgtacta ttgaaaagag cgaagcccct gagtcttctg tcccagttac
     2521 cgaatctaag ggcactacca ccaaagaaac aggtgttact accaaacaaa ccacagccaa
     2581 cccaagtcta accgtatcca cagtcgtccc agtttcatcc tctgcttctt ctcattccgt
     2641 tgtcatcaac agtaacggtg ctaacgtcgt cgttccaggt gctttaggtt tggctggtgt
     2701 tgctatgtta ttcttataac tcgagtggca taaagcaatc ttgatgagga taatgatttt
     2761 tttttgaata tacataaata ctaccgtttt tctgctagat tttgtgatga cgtaaataag
     2821 tacatattac tttttaagcc aagacaagat taagcattaa ctttaccctt ttctttctaa
     2881 gtttcaatat tagttatcac tgtttaaaag ttatggcgag aacgtcggcg gttaaaatat
     2941 attaccctga acggctggat gtgagacgat aaatcgcagc caagtgagtg aatagatgac
     3001 gcaccacggt cagacacggc cacatcgtat ctcacaggag caagcgcgat aggagcactc
     3061 acacatagta cggtgatccg ctgactcctt tgcccaaata agacgtgagc cactagtgca
     3121 ctgcagtaca tcgaggagaa cttctagtat atctacatac ctaatattat tgccttatta
     3181 aaaatggaat cccaacaatt acatcaaaat ccacattctc ttcaaaatca attgtcctgt
     3241 acttccttgt tcatgtgtgt tcaaaaacgt tatatttata ggataattat actctatttc
     3301 tcaacaagta attggttgtt tggccgagcg gtctaaggcg cctgattcaa gaaatatctt
     3361 gaccgcagtt aactgtggga atactcaggt atcgtaagat gcaagagttc gaatctctta
     3421 gcaaccatta tttttttcct caacataacg agaacacaca ggggcgctat cgcacagaat
     3481 caaattcgat gactggaaat tttttgttaa tttcagaggt cgcctgacgc atataccttt
     3541 ttcaactgaa aaattgggag aaaaaggaaa ggtgagagcg ccggaaccgg cttttcatat
     3601 agaatagaga agcgttcatg actaaatgct tgcatcacaa tacttgaagt tgacaatatt
     3661 atttaaggac ctattgtttt ttccaatagg tggttagcaa tcgtcttact ttctaacttt
     3721 tcttaccttt tacatttcag caatatatat atatatattt caaggatata ccattctaat
     3781 gtctgcccct aagaagatcg tcgttttgcc aggtgaccac gttggtcaag aaatcacagc
     3841 cgaagccatt aaggttctta aagctatttc tgatgttcgt tccaatgtca agttcgattt
     3901 cgaaaatcat ttaattggtg gtgctgctat cgatgctaca ggtgttccac ttccagatga
     3961 ggcgctggaa gcctccaaga aggctgatgc cgttttgtta ggtgctgtgg gtggtcctaa
     4021 atggggaacc ggtagtgtta gacctgaaca aggtttacta aaaatccgta aagaacttca
     4081 attgtacgcc aacttaagac catgtaactt tgcatccgac tctcttttag acttatctcc
     4141 aatcaagcca caatttgcta aaggtactga cttcgttgtt gtcagagaat tagtgggagg
     4201 tatttacttt ggtaagagaa aggaggacga tggtgatggt gtcgcttggg atagtgaaca
     4261 atacaccgtt ccagaagtgc aaagaatcac aagaatggcc gctttcatgg ccctacaaca
     4321 tgagccacca ttgcctattt ggtccttgga taaagctaat gttttggcct cttcaagatt
     4381 atggagaaaa actgtggagg aaaccatcaa gaacgaattt cctacattga aggttcaaca
     4441 tcaattgatt gattctgccg ccatgatcct agttaagaac ccaacccacc taaatggtat
     4501 tataatcacc agcaacatgt ttggtgatat catctccgat gaagcctccg ttatcccagg
     4561 ttccttgggt ttgttgccat ctgcgtcctt ggcctctttg ccagacaaga acaccgcatt
     4621 tggtttgtac gaaccatgcc acggttctgc tccagatttg ccaaagaata aggtcaaccc
     4681 tatcgccact atcttgtctg ctgcaatgat gttgaaattg tcattgaact tgcctgaaga
     4741 aggtaaggcc attgaagatg cagttaaaaa ggttttggat gcaggtatca gaactggtga
     4801 tttaggtggt tccaacagta ccaccgaagt cggtgatgct gtcgccgaag aagttaagaa
     4861 aatccttgct taacctggag gacccttctc tttagactat tctactctta tgcacgtaaa
     4921 aaattctagg aaatatgtat taactaggag taaaataacc ggctagtggc attcatatag
     4981 ccgtctgttt acatctacat cacacatttc gagtgtatat ctcgcaacgt tggcgttaaa
     5041 taggcaggag tatcacgtgc tataaaaata attataattt aaatttttta atataaatat
     5101 ataaattaaa aatagaaagt aaaaaaagaa attaaagaaa aaatagtttt tgttttccga
     5161 agatgtaaaa gactctaggg ggatcgccaa caaatactac cttttatctt gctcttcctg
     5221 ctctcaggta ttaatgccga attgtttcat cttgtctgtg tagaaaacca cacacgaaaa
     5281 tcctgtgatt ttacatttta cttatcgtta atcgaatgta tatctattta atctgctttt
     5341 cttgtctaat aaatatatat gtaaagtacg ctttttgttg aaatttttta aacctttgtt
     5401 tatttttttt tcttcattcc gtaactcttc taccttcttt atttactttc taaaatccaa
     5461 atacaaaaca taaaaataaa taaacacaga gtaaattccc aaattattcc atcattaaaa
     5521 gatacgaggc gcgtgtaagt tacaggcaag cgatccgtcc cgagcggccg cgattatcaa
     5581 aaaggatctt cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta
     5641 tatatgagta aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag
     5701 cgatctgtct atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga
     5761 tacgggaggg cttaccatct ggccccagtg ctgcaatgat accgcgggac ccacgctcac
     5821 cggctccaga tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc
     5881 ctgcaacttt atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta
     5941 gttcgccagt taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac
     6001 gctcgtcgtt tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat
     6061 gatcccccat gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa
     6121 gtaagttggc cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg
     6181 tcatgccatc cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag
     6241 aatagtgtat gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc
     6301 cacatagcag aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct
     6361 caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat
     6421 cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg
     6481 ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc
     6541 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     6601 tttagaaaaa taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgtca
     6661 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     6721 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     6781 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     6841 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     6901 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     6961 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     7021 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     7081 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     7141 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     7201 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     7261 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     7321 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     7381 tgttctttcc tgcgttatcc cctgattctg tggataaccg tgcggccgc
//
