LOCUS       sequence-113791-        7404 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Yeast expression plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7404)
  AUTHORS   Chen Y, Partow S, Scalcinati G, Siewers V, Nielsen J
  TITLE     Enhancing the copy number of episomal plasmids in Saccharomyces 
            cerevisiae for improved protein production.
  JOURNAL   FEMS Yeast Res. 2012 Aug;12(5):598-607. doi: 
            10.1111/j.1567-1364.2012.00809.x. Epub 2012 Apr 25.
  PUBMED    22487308
REFERENCE   2  (bases 1 to 7404)
  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"; doi: 
            "10.1111/j.1567-1364.2012.00809.x"; journalName: "FEMS Yeast Res"; 
            date: "2012-08"; volume: "12"; issue: "5"; pages: "598-607"
COMMENT     Sequence Label: pSP-GM1
FEATURES             Location/Qualifiers
     source          1..7404
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(3..25)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     125..144
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        170..390
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             391..1194
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     rep_origin      complement(1325..1780)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1462..1483)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     1674..1693
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     terminator      complement(1885..2050)
                     /gene="S. cerevisiae ADH1"
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae 
                     alcohol dehydrogenase 1 (ADH1) gene"
     primer_bind     1971..1995
                     /label=pHybLex-R
                     /note="ADH terminator, reverse primer"
     CDS             complement(2198..2221)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     promoter        complement(2255..2661)
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     primer_bind     3663..3682
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3663..3681
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3699..3728
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     terminator      3758..3947
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4046..4063)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4217..4805)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4297..4316)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4976..5767,5768..5836))
                     /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     5599..5618
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5837..5941)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      5968..7310
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(6931..6978)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     primer_bind     7351..7369
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc ggatgccggg
       61 agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg ggtgtcgggg ctggcttaac
      121 tatgcggcat cagagcagat tgtactgaga gtgcaccata ccacagcttt tcaattcaat
      181 tcatcatttt ttttttattc ttttttttga tttcggtttc tttgaaattt ttttgattcg
      241 gtaatctccg aacagaagga agaacgaagg aaggagcaca gacttagatt ggtatatata
      301 cgcatatgta gtgttgaaga aacatgaaat tgcccagtat tcttaaccca actgcacaga
      361 acaaaaacct gcaggaaacg aagataaatc atgtcgaaag ctacatataa ggaacgtgct
      421 gctactcatc ctagtcctgt tgctgccaag ctatttaata tcatgcacga aaagcaaaca
      481 aacttgtgtg cttcattgga tgttcgtacc accaaggaat tactggagtt agttgaagca
      541 ttaggtccca aaatttgttt actaaaaaca catgtggata tcttgactga tttttccatg
      601 gagggcacag ttaagccgct aaaggcatta tccgccaagt acaatttttt actcttcgaa
      661 gacagaaaat ttgctgacat tggtaataca gtcaaattgc agtactctgc gggtgtatac
      721 agaatagcag aatgggcaga cattacgaat gcacacggtg tggtgggccc aggtattgtt
      781 agcggtttga agcaggcggc agaagaagta acaaaggaac ctagaggcct tttgatgtta
      841 gcagaattgt catgcaaggg ctccctatct actggagaat atactaaggg tactgttgac
      901 attgcgaaga gcgacaaaga ttttgttatc ggctttattg ctcaaagaga catgggtgga
      961 agagatgaag gttacgattg gttgattatg acacccggtg tgggtttaga tgacaaggga
     1021 gacgcattgg gtcaacagta tagaaccgtg gatgatgtgg tctctacagg atctgacatt
     1081 attattgttg gaagaggact atttgcaaag ggaagggatg ctaaggtaga gggtgaacgt
     1141 tacagaaaag caggctggga agcatatttg agaagatgcg gccagcaaaa ctaaaaaact
     1201 gtattataag taaatgcatg tatactaaac tcacaaatta gagcttcaat ttaattatat
     1261 cagttattac cctatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     1321 caggaaattg taaacgttaa tattttgtta aaattcgcgt taaatttttg ttaaatcagc
     1381 tcatttttta accaataggc cgaaatcggc aaaatccctt ataaatcaaa agaatagacc
     1441 gagatagggt tgagtgttgt tccagtttgg aacaagagtc cactattaaa gaacgtggac
     1501 tccaacgtca aagggcgaaa aaccgtctat cagggcgatg gcccactacg tgaaccatca
     1561 ccctaatcaa gttttttggg gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg
     1621 agcccccgat ttagagcttg acggggaaag ccggcgaacg tggcgagaaa ggaagggaag
     1681 aaagcgaaag gagcgggcgc tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc
     1741 accacacccg ccgcgcttaa tgcgccgcta cagggcgcgt ccattcgcca ttcaggctgc
     1801 gcaactgttg ggaagggcga tcggtgcggg cctcttcgct attacgccag ctggataaag
     1861 gcgcgccaaa cgacctagga attggagcga cctcatgcta tacctgagaa agcaacctga
     1921 cctacaggaa agagttactc aagaataaga attttcgttt taaaacctaa gagtcacttt
     1981 aaaatttgta tacacttatt ttttttataa cttatttaat aataaaaatc ataaatcata
     2041 agaaattcgc ttatttagaa gtgtcaacaa cgtatctacc aacgatttga cccttttcca
     2101 tcttttcgta aatttctggc aaggtagaca agccgacaac cttgattgga gacttgacca
     2161 aacctctggc gaagaattgt taattaagag ctcagatctt atcgtcgtca tccttgtaat
     2221 ccatcgatac tagtgcggcc gcttgtaatt aaaacttaga ttagattgct atgctttctt
     2281 tctaatgagc aagaagtaaa aaaagttgta atagaacaag aaaaatgaaa ctgaaacttg
     2341 agaaattgaa gaccgtttat taacttaaat atcaatggga ggtcatcgaa agagaaaaaa
     2401 atcaaaaaaa aaaattttca agaaaaagaa acgtgataaa aatttttatt gcctttttcg
     2461 acgaagaaaa agaaacgagg cggtctcttt tttcttttcc aaacctttag tacgggtaat
     2521 taacgacacc ctagaggaag aaagagggga aatttagtat gctgtgcttg ggtgttttga
     2581 agtggtacgg cgatgcgcgg agtccgagaa aatctggaag agtaaaaaag gagtagaaac
     2641 attttgaagc tatggtgtgt gcggccggcc tggaagtacc ttcaaagaat ggggtcttat
     2701 cttgttttgc aagtaccact gagcaggata ataatagaaa tgataatata ctatagtaga
     2761 gataacgtcg atgacttccc atactgtaat tgcttttagt tgtgtatttt tagtgtgcaa
     2821 gtttctgtaa atcgattaat ttttttttct ttcctctttt tattaacctt aatttttatt
     2881 ttagattcct gacttcaact caagacgcac agatattata acatctgcat aataggcatt
     2941 tgcaagaatt actcgtgagt aaggaaagag tgaggaacta tcgcatacct gcatttaaag
     3001 atgccgattt gggcgcgaat cctttatttt ggcttcaccc tcatactatt atcagggcca
     3061 gaaaaaggaa gtgtttccct ccttcttgaa ttgatgttac cctcataaag cacgtggcct
     3121 cttatcgaga aagaaattac cgtcgctcgt gatttgtttg caaaaagaac aaaactgaaa
     3181 aaacccagac acgctcgact tcctgtcttc ctattgattg cagcttccaa tttcgtcaca
     3241 caacaaggtc ctagcgacgg ctcacaggtt ttgtaacaag caatcgaagg ttctggaatg
     3301 gcgggaaagg gtttagtacc acatgctatg atgcccactg tgatctccag agcaaagttc
     3361 gttcgatcgt actgttactc tctctctttc aaacagaatt gtccgaatcg tgtgacaaca
     3421 acagcctgtt ctcacacact cttttcttct aaccaagggg gtggtttagt ttagtagaac
     3481 ctcgtgaaac ttacatttac atatatataa acttgcataa attggtcaat gcaagaaata
     3541 catatttggt cttttctaat tcgtagtttt tcaagttctt agatgctttc tttttctctt
     3601 ttttacagat catcaaggaa gtaattatct actttttaca acaaatataa aacaaggatc
     3661 cgtaatacga ctcactatag ggcccgggcg tcgacatgga acagaagttg atttccgaag
     3721 aagacctcga gtaagcttgg taccgcggct agctaagatc cgctctaacc gaaaaggaag
     3781 gagttagaca acctgaagtc taggtcccta tttatttttt tatagttatg ttagtattaa
     3841 gaacgttatt tatatttcaa atttttcttt tttttctgta cagacgcgtg tacgcatgta
     3901 acattatact gaaaaccttg cttgagaagg ttttgggacg ctcgaagatc ctccggatcg
     3961 tttcgccggc gtttatccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     4021 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     4081 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     4141 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     4201 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     4261 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     4321 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     4381 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
     4441 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     4501 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     4561 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     4621 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     4681 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     4741 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     4801 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     4861 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     4921 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     4981 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     5041 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     5101 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     5161 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     5221 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     5281 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     5341 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     5401 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     5461 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     5521 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     5581 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
     5641 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
     5701 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
     5761 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
     5821 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
     5881 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
     5941 gcacatttcc ccgaaaagtg ccacctgaac gaagcatctg tgcttcattt tgtagaacaa
     6001 aaatgcaacg cgagagcgct aatttttcaa acaaagaatc tgagctgcat ttttacagaa
     6061 cagaaatgca acgcgaaagc gctattttac caacgaagaa tctgtgcttc atttttgtaa
     6121 aacaaaaatg caacgcgaga gcgctaattt ttcaaacaaa gaatctgagc tgcattttta
     6181 cagaacagaa atgcaacgcg agagcgctat tttaccaaca aagaatctat acttcttttt
     6241 tgttctacaa aaatgcatcc cgagagcgct atttttctaa caaagcatct tagattactt
     6301 tttttctcct ttgtgcgctc tataatgcag tctcttgata actttttgca ctgtaggtcc
     6361 gttaaggtta gaagaaggct actttggtgt ctattttctc ttccataaaa aaagcctgac
     6421 tccacttccc gcgtttactg attactagcg aagctgcggg tgcatttttt caagataaag
     6481 gcatccccga ttatattcta taccgatgtg gattgcgcat actttgtgaa cagaaagtga
     6541 tagcgttgat gattcttcat tggtcagaaa attatgaacg gtttcttcta ttttgtctct
     6601 atatactacg tataggaaat gtttacattt tcgtattgtt ttcgattcac tctatgaata
     6661 gttcttacta caattttttt gtctaaagag taatactaga gataaacata aaaaatgtag
     6721 aggtcgagtt tagatgcaag ttcaaggagc gaaaggtgga tgggtaggtt atatagggat
     6781 atagcacaga gatatatagc aaagagatac ttttgagcaa tgtttgtgga agcggtattc
     6841 gcaatatttt agtagctcgt tacagtccgg tgcgtttttg gttttttgaa agtgcgtctt
     6901 cagagcgctt ttggttttca aaagcgctct gaagttccta tactttctag agaataggaa
     6961 cttcggaata ggaacttcaa agcgtttccg aaaacgagcg cttccgaaaa tgcaacgcga
     7021 gctgcgcaca tacagctcac tgttcacgtc gcacctatat ctgcgtgttg cctgtatata
     7081 tatatacatg agaagaacgg catagtgcgt gtttatgctt aaatgcgtac ttatatgcgt
     7141 ctatttatgt aggatgaaag gtagtctagt acctcctgtg atattatccc attccatgcg
     7201 gggtatcgta tgcttccttc agcactaccc tttagctgtt ctatatgctg ccactcctca
     7261 attggattag tctcatcctt caatgctatc atttcctttg atattggatc atactaagaa
     7321 accattatta tcatgacatt aacctataaa aataggcgta tcacgaggcc ctttcgtctc
     7381 gcgcgtttcg gtgatgacgg tgaa
//
