LOCUS       sequence-64877-e        5143 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5143)
  AUTHORS   Shen MW, Fang F, Sandmeyer S, Da Silva NA
  TITLE     Development and characterization of a vector set with regulated 
            promoters for systematic metabolic engineering in Saccharomyces 
            cerevisiae.
  JOURNAL   Yeast. 2012 Dec;29(12):495-503. doi: 10.1002/yea.2930. Epub 2012 Nov
            19.
  PUBMED    23166051
REFERENCE   2  (bases 1 to 5143)
  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: "10.1002/yea.2930"; 
            journalName: "Yeast"; date: "2012-12"; volume: "29"; issue: "12"; 
            pages: "495-503"
COMMENT     Sequence Label: pXP712
FEATURES             Location/Qualifiers
     source          1..5143
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      734..981
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(750..768)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    complement(1027..1060)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        1078..1294
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             1295..2098
                     /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
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLATGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     protein_bind    complement(2177..2210)
                     /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(2740..2756)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2740..2756)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2753..2775)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2764..2780
                     /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(2788..2794,2795..2812,2813..2818))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2833..2854
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2971..2988)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3142..3730)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3222..3241)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3901..4692,4693..4761))
                     /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     4524..4543
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4762..4866)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4934..4952
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4990..5012)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5112..5131
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 catatgaggc atttgcaaga attactcgtg agtaaggaaa gagtgaggaa ctatcgcata
       61 cctgcattta aagatgccga tttgggcgcg aatcctttat tttggcttca ccctcatact
      121 attatcaggg ccagaaaaag gaagtgtttc cctccttctt gaattgatgt taccctcata
      181 aagcacgtgg cctcttatcg agaaagaaat taccgtcgct cgtgatttgt ttgcaaaaag
      241 aacaaaactg aaaaaaccca gacacgctcg acttcctgtc ttcctattga ttgcagcttc
      301 caatttcgtc acacaacaag gtcctagcga cggctcacag gttttgtaac aagcaatcga
      361 aggttctgga atggcgggaa agggtttagt accacatgct atgatgccca ctgtgatctc
      421 cagagcaaag ttcgttcgat cgtactgtta ctctctctct ttcaaacaga attgtccgaa
      481 tcgtgtgaca acaacagcct gttctcacac actcttttct tctaaccaag ggggtggttt
      541 agtttagtag aacctcgtga aacttacatt tacatatata taaacttgca taaattggtc
      601 aatgcaagaa atacatattt ggtcttttct aattcgtagt ttttcaagtt cttagatgct
      661 ttctttttct cttttttaca gatcatcaag gaagtaatta tctacttttt acaacaaata
      721 tactagtctc gagtcatgta attagttatg tcacgcttac attcacgccc tccccccaca
      781 tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt
      841 tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg
      901 tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga
      961 cgctcgaagg ctttaatttg cggccaagct tgcatgcctg caggtcgact ctagaggatc
     1021 cccgggataa cttcgtatag catacattat acgaagttat cagggtccat aaagcttttc
     1081 aattcatctt tttttttttt gttctttttt ttgattccgg tttctttgaa atttttttga
     1141 ttcggtaatc tccgagcaga aggaagaacg aaggaaggag cacagactta gattggtata
     1201 tatacgcata tgtggtgttg aagaaacatg aaattgccca gtattcttaa cccaactgca
     1261 cagaacaaaa acctgcagga aacgaagata aatcatgtcg aaagctacat ataaggaacg
     1321 tgctgctact catcctagtc ctgttgctgc caagctattt aatatcatgc acgaaaagca
     1381 aacaaacttg tgtgcttcat tggatgttcg taccaccaag gaattactgg agttagttga
     1441 agcattaggt cccaaaattt gtttactaaa aacacatgtg gatatcttga ctgatttttc
     1501 catggagggc acagttaagc cgctaaaggc attatccgcc aagtacaatt ttttactctt
     1561 cgaagacaga aaatttgctg acattggtaa tacagtcaaa ttgcagtact ctgcgggtgt
     1621 atacagaata gcagaatggg cagacattac gaatgcacac ggtgtggtgg gcccaggtat
     1681 tgttagcggt ttgaagcagg cggcggaaga agtaacaaag gaacctagag gccttttgat
     1741 gttagcagaa ttgtcatgca agggctccct agctactgga gaatatacta agggtactgt
     1801 tgacattgcg aagagcgaca aagattttgt tatcggcttt attgctcaaa gagacatggg
     1861 tggaagagat gaaggttacg attggttgat tatgacaccc ggtgtgggtt tagatgacaa
     1921 gggagacgca ttgggtcaac agtatagaac cgtggatgat gtggtctcta caggatctga
     1981 cattattatt gttggaagag gactatttgc aaagggaagg gatgctaagg tagagggtga
     2041 acgttacaga aaagcaggct gggaagcata tttgagaaga tgcggccagc aaaactaaaa
     2101 aactgtatta taagtaaatg catgtatact aaactcacaa attagagctt caatttaatt
     2161 atatcagtta ttacccataa cttcgtatag catacattat acgaagttat cccgggtacc
     2221 gagctcgaat tcatcacgtg ctataaaaat aattataatt taaatttttt aatataaata
     2281 tataaattaa aaatagaaag taaaaaaaga aattaaagaa aaaatagttt ttgttttccg
     2341 aagatgtaaa agactctagg gggatcgcca acaaatacta ccttttatct tgctcttcct
     2401 gctctcaggt attaatgccg aattgtttca tcttgtctgt gtagaagacc acacacgaaa
     2461 atcctgtgat tttacatttt acttatcgtt aatcgaatgt atatctattt aatctgcttt
     2521 tcttgtctaa taaatatata tgtaaagtac gctttttgtt gaaatttttt aaacctttgt
     2581 ttattttttt ttcttcattc cgtaactctt ctaccttctt tatttacttt ctaaaatcca
     2641 aatacaaaac ataaaaataa ataaacacag agtaaattcc caaattattc catcattaaa
     2701 agatacgagg cgcgtgtaag ttacgaattc gtaatcatgg tcatagctgt ttcctgtgtg
     2761 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
     2821 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
     2881 ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg
     2941 cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt
     3001 tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat ccacagaatc
     3061 aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa
     3121 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     3181 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     3241 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     3301 cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta ggtatctcag
     3361 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     3421 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     3481 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     3541 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     3601 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     3661 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3721 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3781 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3841 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3901 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3961 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     4021 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     4081 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     4141 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     4201 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     4261 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     4321 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     4381 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     4441 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     4501 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     4561 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     4621 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     4681 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4741 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4801 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4861 tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa gaaaccatta ttatcatgac
     4921 attaacctat aaaaataggc gtatcacgag gccctttcgt ctcgcgcgtt tcggtgatga
     4981 cggtgaaaac ctctgacaca tgcagctccc ggagacggtc acagcttgtc tgtaagcgga
     5041 tgccgggagc agacaagccc gtcagggcgc gtcagcgggt gttggcgggt gtcggggctg
     5101 gcttaactat gcggcatcag agcagattgt actgagagtg cac
//
