LOCUS       sequence-430980-        7632 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual-species protein expression vector for high-level and inducible 
            protein expression of mNeonGreen (codon optimized for yeast) in both
            yeast and bacteria from the same construct..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7632)
  AUTHORS   Waneskog M, Rasmussen TB, Jensen ED
  TITLE     A strategy for successful dual-species protein expression of genes 
            with non-optimal codon usage destined for bacterial and yeast cell 
            factories.
  JOURNAL   Biotechnol Prog. 2024 May 17:e3482. doi: 10.1002/btpr.3482.
  PUBMED    38757558
REFERENCE   2  (bases 1 to 7632)
  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: "Biotechnol 
            Prog. 2024 May 17:e3482. doi: 10.1002/btpr.3482."
COMMENT     Sequence Label: pYB-Dual(mNG(yeast-opt))
FEATURES             Location/Qualifiers
     source          1..7632
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(424..1518)
                     /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"
     promoter        complement(1519..1926)
                     /gene="S. cerevisiae LEU2"
                     /label=LEU2 promoter
     rep_origin      complement(2227..2682)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2364..2385)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2576..2595
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2808..2830
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2822..2839
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2823..2839
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2849..2867
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     terminator      complement(2896..3061)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     2982..3006
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     promoter        3192..3633
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    3192..3309
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     3604..3627
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     promoter        3654..3672
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4618..4638)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(4618..4636)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4657..4673)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4657..4673)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4670..4692)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4681..4697
                     /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)."
     promoter        complement(join(4705..4711,4712..4729,4730..4735))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4750..4771
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4888..4905)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5059..5647)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5139..5158)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5818..6609,6610..6678))
                     /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     6441..6460
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6679..6783)
                     /gene="bla"
                     /label=AmpR promoter
     misc_feature    6820..7323
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     primer_bind     7365..7383
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(7421..7443)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     7543..7562
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 caccattatg ggaaatggtt caagaaggta ttgacttaaa ctccatcaaa tggtcaggtc
       61 attgagtgtt ttttatttgt tgtatttttt tttttttaga gaaaatcctc caatatataa
      121 attaggaatc atagtttcat gattttctgt tacacctaac tttttgtgtg gtgccctcct
      181 ccttgtcaat attaatgtta aagtgcaatt ctttttcctt atcacgttga gccattagta
      241 tcaatttgct tacctgtatt cctttacatc ctcctttttc tccttcttga taaatgtatg
      301 tagattgcgt atatagtttc gtctacccta tgaacatatt ccattttgta atttcgtgtc
      361 gtttctatta tgaatttcat ttataaagtt tatgtacaaa tatcataaaa aaagagaatc
      421 tttttaagca aggattttct taacttcttc ggcgacagca tcaccgactt cggtggtact
      481 gttggaacca cctaaatcac cagttctgat acctgcatcc aaaacctttt taactgcatc
      541 ttcaatggcc ttaccttctt caggcaagtt caatgacaat ttcaacatca ttgcagcaga
      601 caagatagtg gcgatagggt tgaccttatt ctttggcaaa tctggagcag aaccgtggca
      661 tggttcgtac aaaccaaatg cggtgttctt gtctggcaaa gaggccaagg acgcagatgg
      721 caacaaaccc aaggaacctg ggataacgga ggcttcatcg gagatgatat caccaaacat
      781 gttgctggtg attataatac catttaggtg ggttgggttc ttaactagga tcatggcggc
      841 agaatcaatc aattgatgtt gaaccttcaa tgtagggaat tcgttcttga tggtttcctc
      901 cacagttttt ctccataatc ttgaagaggc caaaacatta gctttatcca aggaccaaat
      961 aggcaatggt ggctcatgtt gtagggccat gaaagcggcc attcttgtga ttctttgcac
     1021 ttctggaacg gtgtattgtt cactatccca agcgacacca tcaccatcgt cttcctttct
     1081 cttaccaaag taaatacctc ccactaattc tctgacaaca acgaagtcag tacctttagc
     1141 aaattgtggc ttgattggag ataagtctaa aagagagtcg gatgcaaagt tacatggtct
     1201 taagttggcg tacaattgaa gttctttacg gatttttagt aaaccttgtt caggtctaac
     1261 actaccggta ccccatttag gaccacccac agcacctaac aaaacggcat cagccttctt
     1321 ggaggcttcc agcgcctcat ctggaagtgg aacacctgta gcatcgatag cagcaccacc
     1381 aattaaatga ttttcgaaat cgaacttgac attggaacga acatcagaaa tagctttaag
     1441 aaccttaatg gcttcggctg tgatttcttg accaacgtgg tcacctggca aaacgacgat
     1501 cttcttaggg gcagacatta gaatggtata tccttgaaat atatatatat atattgctga
     1561 aatgtaaaag gtaagaaaag ttagaaagta agacgattgc taaccaccta ttggaaaaaa
     1621 caataggtcc ttaaataata ttgtcaactt caagtattgt gatgcaagca tttagtcatg
     1681 aacgcttctc tattctatat gaaaagccgg ttccggcgct ctcacctttc ctttttctcc
     1741 caatttttca gttgaaaaag gtatatgcgt caggcgacct ctgaaattaa caaaaaattt
     1801 ccagtcatcg aatttgattc tgtgcgatag cgcccctgtg tgttctcgtt atgttgagga
     1861 aaaaaataat ggttgctaag agattcgaac tcttgcatct tacgatacct gagtattccc
     1921 acagttaact gcggtcaaga tatttcttga atcaggcgcc ttagaccgct cggccaaaca
     1981 accaattact tgttgagaaa tagagtataa ttatcctata aatataacgt ttttgaacac
     2041 acatgaacaa ggaagtacag gacaattgat tttgaagaga atgtggattt tgatgtaatt
     2101 gttgggattc catttttaat aaggcaataa tattaggtat gtagatatac tagaagttct
     2161 cctcgaccgg tcgatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc
     2221 atcaggaaat tgtaagcgtt aatattttgt taaaattcgc gttaaatttt tgttaaatca
     2281 gctcattttt taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga
     2341 ccgagatagg gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg
     2401 actccaacgt caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat
     2461 caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag
     2521 ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga
     2581 agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa
     2641 ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtccattcgc cattcaggct
     2701 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     2761 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     2821 ttgtaaaacg acggccagtg agcgcgcgta atacgactca ctatagggcg aattgggtac
     2881 cgggcccccc ctcgagagcg acctcatgct atacctgaga aagcaacctg acctacagga
     2941 aagagttact caagaataag aattttcgtt ttaaaaccta agagtcactt taaaatttgt
     3001 atacacttat tttttttata acttatttaa taataaaaat cataaatcat aagaaattcg
     3061 cttatttaga agtgtcaaca acgtatctac caacggaatg cgtgcgatga gccccattat
     3121 cttagcctaa aaaaaccttc tctttggaac tttcagtaat acgcttaact gctcattgct
     3181 atattgaagt acggattaga agccgccgag cgggcgacag ccctccgacg gaagactctc
     3241 ctccgtgcgt cctggtcttc accggtcgcg ttcctgaaac gcagatgtgc ctcgcgccgc
     3301 actgctccga acaataaaga ttctacaata ctagctttta tggttatgaa gaggaaaaat
     3361 tggcagtaac ctggccccac aaaccttcaa atcaacgaat caaattaaca accataggat
     3421 aataatgcga ttagtttttt agccttattt ctggggtaat taatcagcga agcgatgatt
     3481 tttgatctat taacagatat ataaatgcaa aagctgcata accactttaa ctaatacttt
     3541 caacattttc ggtttgtatt acttcttatt caaatgtcat aaaagtatca acaaaaaatt
     3601 gttaatatac ctctatactt taacgtcaag gagaaaaaac tataatctgt cattaatacg
     3661 actcactata ggtctagaga aagaggggag aattaaaaat ggtgtctaaa ggtgaagagg
     3721 acaacatggc ttctttgcca gctactcatg aattgcatat tttcggttcc atcaacggtg
     3781 ttgatttcga tatggttggt caaggtactg gtaatccaaa tgatggttac gaagagttga
     3841 acctgaaatc tacaaaaggt gacttgcaat tctccccatg gattttggtt ccacatattg
     3901 gttacggttt ccaccaatat ttgccatatc cagatggtat gtctccattt caagctgcta
     3961 tggttgatgg ttctggttac caagttcata gaaccatgca atttgaagat ggtgcttctt
     4021 tgaccgtcaa ctacagatat acttacgaag gttcccatat taagggtgaa gctcaagtaa
     4081 aaggtacagg ttttccagct gatggtccag ttatgactaa ttctttgact gctgctgatt
     4141 ggtgcagatc caaaaagact tacccaaacg ataagaccat catctctact ttcaagtggt
     4201 cttacactac tggtaacggt aagagatata gatctactgc tagaactact tacaccttcg
     4261 ctaaacctat ggctgctaac tacttgaaga atcaacctat gtacgtgttc agaaagaccg
     4321 aattgaaaca ctccaaaacc gagttgaact tcaaagaatg gcaaaaggct ttcaccgatg
     4381 ttatgggtat ggatgaatta tacaaataag gatccatcgc gtgcattcat ccgctctaac
     4441 cgaaaaggaa ggagttagac aacctgaagt ctaggtccct atttattttt ttatagttat
     4501 gttagtatta agaacgttat ttatatttca aatttttctt ttttttctgt acagacgcgt
     4561 gtacgcatgt aacattatac tgaaaacctt gcttgagaag agctccagct tttgttccct
     4621 ttagtgaggg ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
     4681 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
     4741 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
     4801 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
     4861 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
     4921 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     4981 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     5041 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     5101 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     5161 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     5221 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     5281 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     5341 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     5401 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     5461 gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc
     5521 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     5581 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     5641 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     5701 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     5761 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     5821 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     5881 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     5941 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     6001 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     6061 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     6121 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     6181 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     6241 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     6301 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     6361 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     6421 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     6481 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     6541 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     6601 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     6661 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     6721 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     6781 gcgcacattt ccccgaaaag tgccacctgg gtccttttca tcacgtgcta taaaaataat
     6841 tataatttaa attttttaat ataaatatat aaattaaaaa tagaaagtaa aaaaagaaat
     6901 taaagaaaaa atagtttttg ttttccgaag atgtaaaaga ctctaggggg atcgccaaca
     6961 aatactacct tttatcttgc tcttcctgct ctcaggtatt aatgccgaat tgtttcatct
     7021 tgtctgtgta gaagaccaca cacgaaaatc ctgtgatttt acattttact tatcgttaat
     7081 cgaatgtata tctatttaat ctgcttttct tgtctaataa atatatatgt aaagtacgct
     7141 ttttgttgaa attttttaaa cctttgttta tttttttttc ttcattccgt aactcttcta
     7201 ccttctttat ttactttcta aaatccaaat acaaaacata aaaataaata aacacagagt
     7261 aaattcccaa attattccat cattaaaaga tacgaggcgc gtgtaagtta caggcaagcg
     7321 atccgtccta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     7381 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     7441 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     7501 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
     7561 tactgagagt gcaccatatc gactacgtcg ttaaggccgt ttctgacaga gtaaaattct
     7621 tgagggaact tt
//
