LOCUS       sequence-133491-        6980 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast chr. integration vector with DS3 selection marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6980)
  AUTHORS   Rugbjerg P, Myling-Petersen N, Sommer MO
  TITLE     Flexible metabolic pathway construction using modular and divisible 
            selection gene regulators.
  JOURNAL   Metab Eng. 2015 Sep;31:189-97. doi: 10.1016/j.ymben.2015.08.004. 
            Epub 2015 Aug 21.
  PUBMED    26303342
REFERENCE   2  (bases 1 to 6980)
  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: "2015-09"; volume: "31:189-97. doi"; pages: " 
            10.1016/j.ymben.2015.08.004. Epub 2015 Aug 21"
COMMENT     Sequence Label: pDS3-X4
FEATURES             Location/Qualifiers
     source          1..6980
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    208..241
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     terminator      complement(842..1007)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     928..952
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(1058..1771)
                     /codon_start=1
                     /product="S65T variant of Aequorea victoria green 
                     fluorescent protein (Heim et al., 1995)"
                     /label=GFP (S65T)
                     /note="excitable with blue light"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKRHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(1098..1118)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     1713..1741
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     promoter        complement(1799..2205)
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     protein_bind    2444..2477
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             2883..2903
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2916..3149
                     /codon_start=1
                     /gene="UL48"
                     /product="transcriptional activation domain of herpes 
                     simplex virus protein VP16 (Triezenberg et al., 1988; 
                     Cousens et al., 1989)"
                     /label=VP16 AD
                     /translation="APPTDVSLGDELHLDGEDVAMAHADALDDFDLDMLGDGDSPGPGF
                     TPHDSAPYGALDMADFEFEQMFTDALGIDEYGG"
     protein_bind    3159..3183
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    3767..3800
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(3843..3862)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3844..3862)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3872..3888)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3881..3903)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     protein_bind    complement(4442..4475)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     primer_bind     complement(4945..4961)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5060..5077)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5231..5819)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5311..5330)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5943..6734,6735..6803))
                     /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     6566..6585
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6804..6895)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     6937..6956
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     join(6962..6980,1)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
ORIGIN
        1 gtttgcctgg cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga
       61 aacgccgtag cgccgatggt agtgtgggga ctccccatgc gagagtaggg aactgccagg
      121 catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgcccggg ctaattatgg
      181 ggtgtcgccc ttattcgact ctatagtgaa gttcctattc tctagaaagt ataggaactt
      241 ctgaagtggg gatttaaatg cggccgcgct gagggtttaa tcccaaagct aagagtccca
      301 ttttattctt ctatatgtat attttcgata ctctaaacca ccctacaatg tagccctata
      361 ctaaatctgc tcaattttca gcttctacaa gtgactcgag accacgtgga aagatccaac
      421 tactccagca caacgattca atataatcga ttgctccact cataagaggc aagaacaagc
      481 ttcaactttt ggtaagccgc cgtttataaa cagggaagat gtcctttgtc aagggaggca
      541 cagagcatgg ccaatttggc aaattgcagg tttttctgag tgaaaaatga aaaagcattg
      601 tagtagagtc ggctcactga aaaaccgggg aggacgaaaa ggtttccagc cacagttgta
      661 gtcacgtgcg cgccatgctg actaatggca gccgtcgttg ggcagaagag aattagtatg
      721 gtacaggata cgctaattgc gctccaacta ccaaggttgt tgagggaaca ctggggcaat
      781 aggctgtcgc cattcaagag cagattaagt cctcagcgag ctcgcatgga atgcgtgcga
      841 tgagcgacct catgctatac ctgagaaagc aacctgacct acaggaaaga gttactcaag
      901 aataagaatt ttcgttttaa aacctaagag tcactttaaa atttgtatac acttattttt
      961 tttataactt atttaataat aaaaatcata aatcataaga aattcgctta tttagaagtg
     1021 tcaacaacgt atctaccaac ggaatgcgtg cgattcattt gtatagttca tccatgccat
     1081 gtgtaatccc agcagctgtt acaaactcaa gaaggaccat gtggtctctc ttttcgttgg
     1141 gatctttcga aagggcagat tgataggaca ggtaatggtt gtctggtaaa aggacagggc
     1201 catcgccaat tggagtattt tgttgataat ggtctgctag ttgaacgctt ccatcttcaa
     1261 tgttgtgtct aattttgaag ttaactttga ttccattctt ttgtttgtct gccatgatgt
     1321 atacattgtg tgagttatag ttgtattcca atttgtgtcc aagaatgttt ccatcttctt
     1381 taaaatcaat accttttaac tcgattctat taacaagggt atcaccttca aacttgactt
     1441 cagcacgtgt cttgtagttc ccgtcatctt tgaaaaatat agttctttcc tgtacataac
     1501 cttcgggcat ggcactcttg aaaaagtcat gccgtttcat atgatctggg tatcttgcaa
     1561 agcattgaag accataaccg aaagtagtga caagtgttgg ccatggaaca ggtagttttc
     1621 cagtagtgca aataaattta agggtaagtt ttccgtatgt tgcatcacct tcaccctctc
     1681 cactgacaga aaatttgtgc ccattaacat caccatctaa ttcaacaaga attgggacaa
     1741 ctccagtgaa aagttcttct cctttactca ttgttttacc tgcactttgt aattaaaact
     1801 tagattagat tgctatgctt tctttctaat gagcaagaag taaaaaaagt tgtaatagaa
     1861 caagaaaaat gaaactgaaa cttgagaaat tgaagaccgt ttattaactt aaatatcaat
     1921 gggaggtcat cgaaagagaa aaaaatcaaa aaaaaaaatt ttcaagaaaa agaaacgtga
     1981 taaaaatttt tattgccttt ttcgacgaag aaaaagaaac gaggcggtct cttttttctt
     2041 ttccaaacct ttagtacggg taattaacga caccctagag gaagaaagag gggaaattta
     2101 gtatgctgtg cttgggtgtt ttgaagtggt acggcgatgc gcggagtccg agaaaatctg
     2161 gaagagtaaa aaaggagtag aaacattttg aagctatggt gtgtgcatcg cgtgcattca
     2221 tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt
     2281 tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg
     2341 tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga
     2401 cgctccgaag atcgcgtcat aacagctttc gacaaccctt aatataactt cgtataatgt
     2461 atgctatacg aagttattag gtctagagat ctgtttagct tgcctcgtcc ccgccgggtc
     2521 acccggccag cgacatggag gcccagaata ccctccttga cagtcttgac gtgcgcagct
     2581 caggggcatg atgtgactgt cgcccgtaca tttagcccat acatccccat gtataatcat
     2641 ttgcatccat acattttgat ggccgcacgg cgcgaagcaa aaattacggc tcctcgctgc
     2701 agacctgcga gcagggaaac gctcccctca cagacgcgtt gaattgtccc cacgccgcgc
     2761 ccctgtagag aaatataaaa ggttaggatt tgccactgag gttcttcttt catatacttc
     2821 cttttaaaat cttgctagga tacagttctc acatcacatc cgaacataaa caagaacata
     2881 tgcccaagaa gaagcggaag gtcaagcttt cgacggcccc cccgaccgat gtcagcctgg
     2941 gggacgagct ccacttagac ggcgaggacg tggcgatggc gcatgccgac gcgctagacg
     3001 atttcgatct ggacatgttg ggggacgggg attccccggg gccgggattt accccccacg
     3061 actccgcccc ctacggcgct ctggatatgg ccgacttcga gtttgagcag atgtttaccg
     3121 atgcccttgg aattgacgag tacggtgggt cgaatcaaac aagtttgtac aaaaaagcag
     3181 gctctgcggc cgcccccttc acctccagct cctcactgcc actctacaac cagcaggtgg
     3241 gcgactgctg catcatcagg gtcagcctgg atgtggacaa cggcaacatg tacaagagca
     3301 tcctggtgac cagccaggat aaggctccga ctgtcatccg gaaagccatg gacaaacaca
     3361 acctagatga ggacgagccc gaggactatg agctgctgca gatcatctca gaggaccaca
     3421 agctgaaaat tccagagaac gccaatgtgt tctatgccat gaactctgcc gccaactatg
     3481 acttcatcct aaagaagcgg taactcgagg acaataaaaa gattcttgtt ttcaagaact
     3541 tgtcatttgt atagtttttt tatattgtag ttgttctatt ttaatcaaat gttagcgtga
     3601 tttatatttt ttttcgcctc gacatcatct gcccagatgc gaagttaagt gcgcagaaag
     3661 taatatcatg cgtcaatcgt atgtgaatgc tggtcgctat actgctgtcg attcgatact
     3721 aacgccgcca tccagtgtcg aaaacgagct ctcgagaacc cttaatataa cttcgtataa
     3781 tgtatgctat acgaagttat taggtgatat cagatccact agtggcctat gcacccaatt
     3841 cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa cgtcgtgact
     3901 gggaaaaccc tggcgttacc cctgcaggac tagtgctgag gcattaataa caggcatggg
     3961 aagattcgct tttttttttt gaattacaat agtatgtctg atgtctgcaa gaagtaacag
     4021 gcgtgtgcac aagaatacgt gtgtgtgcgt aagcgtatgc actggtggca taacttatct
     4081 aagaagtata tatcactgac atagaaatgt agatatacag gtatttttct cgataatcga
     4141 taaaaatctc gtcgcgctga accaaacttg gtggttacgg agagtttttc tctcatcatt
     4201 actgtctttc gcattgattt cccctttgac cgataaaatc ccttggattc ataagattaa
     4261 acaaagaggt gatcaaagag aaccctgtga aagtttatgt ttataaccgg gcataaagtg
     4321 aactagacac tttcaagaag ccaaccaaag catgagtaac gaagcttacc agcatgatca
     4381 taccgtaaat cctcaccaga ttaagacctc agcgcggccg caaatttaaa taaaatgaag
     4441 tgaagttcct atactttcta gagaatagga acttctatag tgagtcgaat aagggcgaca
     4501 caaaatttat tctaaatgca taataaatac tgataacatc ttatagtttg tattatattt
     4561 tgtattatcg ttgacatgta taattttgat atcaaaaact gattttccct ttattatttt
     4621 cgagatttat tttcttaatt ctctttaaca aactagaaat attgtatata caaaaaatca
     4681 taaataatag atgaatagtt taattatagg tgttcatcaa tcgaaaaagc aacgtatctt
     4741 atttaaagtg cgttgctttt ttctcattta taaggttaaa taattctcat atatcaagca
     4801 aagtgacagg cgcccttaaa tattctgaca aatgctcttt ccctaaactc cccccataaa
     4861 aaaacccgcc gaagcgggtt tttacgttat ttgcggatta acgattactc gttatcagaa
     4921 ccgcccaggg ggcccgagct taagactggc cgtcgtttta caacacagaa agagtttgta
     4981 gaaacgcaaa aaggccatcc gtcaggggcc ttctgcttag tttgatgcct ggcagttccc
     5041 tactctcgcc ttccgcttcc tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc
     5101 gagcggtatc agctcactca aaggcggtaa tacggttatc cacagaatca ggggataacg
     5161 caggaaagaa catgtgagca aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt
     5221 tgctggcgtt tttccatagg ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa
     5281 gtcagaggtg gcgaaacccg acaggactat aaagatacca ggcgtttccc cctggaagct
     5341 ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc
     5401 cttcgggaag cgtggcgctt tctcatagct cacgctgtag gtatctcagt tcggtgtagg
     5461 tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct
     5521 tatccggtaa ctatcgtctt gagtccaacc cggtaagaca cgacttatcg ccactggcag
     5581 cagccactgg taacaggatt agcagagcga ggtatgtagg cggtgctaca gagttcttga
     5641 agtggtgggc taactacggc tacactagaa gaacagtatt tggtatctgc gctctgctga
     5701 agccagttac cttcggaaaa agagttggta gctcttgatc cggcaaacaa accaccgctg
     5761 gtagcggtgg tttttttgtt tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag
     5821 aagatccttt gatcttttct acggggtctg acgctcagtg gaacgacgcg cgcgtaactc
     5881 acgttaaggg attttggtca tgagcttgcg ccgtcccgtc aagtcagcgt aatgctctgc
     5941 ttttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     6001 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     6061 cccagcgctg cgatgatacc gcgagaacca cgctcaccgg ctccggattt atcagcaata
     6121 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     6181 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     6241 aacgttgttg ccatcgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     6301 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     6361 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     6421 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     6481 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     6541 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     6601 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     6661 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     6721 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     6781 acacggaaat gttgaatact catattcttc ctttttcaat attattgaag catttatcag
     6841 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     6901 gtcagtgtta caaccaatta accaattctg aacattatcg cgagcccatt tatacctgaa
     6961 tatggctcat aacacccctt
//
