LOCUS       sequence-64882-e        4734 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 4734)
  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 4734)
  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: pXP732
FEATURES             Location/Qualifiers
     source          1..4734
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        22..256
                     /gene="S. cerevisiae CUP1"
                     /label=CUP1 promoter
                     /note="inducible metallothionein promoter, regulated by 
                     Cu2+"
     terminator      325..572
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(341..359)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    complement(618..651)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        669..885
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             886..1689
                     /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(1768..1801)
                     /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(2331..2347)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2331..2347)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2344..2366)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2355..2371
                     /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(2379..2385,2386..2403,2404..2409))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2424..2445
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2562..2579)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2733..3321)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2813..2832)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3492..4283,4284..4352))
                     /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     4115..4134
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4353..4457)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4525..4543
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4581..4603)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4703..4722
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 catatgatct tagccttgtt actagttaga aaaagacatt tttgctgtca gtcactgtca
       61 agagattctt ttgctggcat ttcttctaga agcaaaaaga gcgatgcgtc ttttccgctg
      121 aaccgttcca gcaaaaaaga ctaccaacgc aatatggatt gtcagaatca tataaaagag
      181 aagcaaataa ctccttgtct tgtatcaatt gcattataat atcttcttgt tagtgcaata
      241 tcatatagaa gtcatcgaaa tagatattaa gaaaaacaaa ctgtacaatc aatcaatcaa
      301 tcatcacata aacctaggct cgagtcatgt aattagttat gtcacgctta cattcacgcc
      361 ctccccccac atccgctcta accgaaaagg aaggagttag acaacctgaa gtctaggtcc
      421 ctatttattt ttttatagtt atgttagtat taagaacgtt atttatattt caaatttttc
      481 ttttttttct gtacagacgc gtgtacgcat gtaacattat actgaaaacc ttgcttgaga
      541 aggttttggg acgctcgaag gctttaattt gcggccaagc ttgcatgcct gcaggtcgac
      601 tctagaggat ccccgggata acttcgtata gcatacatta tacgaagtta tcagggtcca
      661 taaagctttt caattcatct tttttttttt tgttcttttt tttgattccg gtttctttga
      721 aatttttttg attcggtaat ctccgagcag aaggaagaac gaaggaagga gcacagactt
      781 agattggtat atatacgcat atgtggtgtt gaagaaacat gaaattgccc agtattctta
      841 acccaactgc acagaacaaa aacctgcagg aaacgaagat aaatcatgtc gaaagctaca
      901 tataaggaac gtgctgctac tcatcctagt cctgttgctg ccaagctatt taatatcatg
      961 cacgaaaagc aaacaaactt gtgtgcttca ttggatgttc gtaccaccaa ggaattactg
     1021 gagttagttg aagcattagg tcccaaaatt tgtttactaa aaacacatgt ggatatcttg
     1081 actgattttt ccatggaggg cacagttaag ccgctaaagg cattatccgc caagtacaat
     1141 tttttactct tcgaagacag aaaatttgct gacattggta atacagtcaa attgcagtac
     1201 tctgcgggtg tatacagaat agcagaatgg gcagacatta cgaatgcaca cggtgtggtg
     1261 ggcccaggta ttgttagcgg tttgaagcag gcggcggaag aagtaacaaa ggaacctaga
     1321 ggccttttga tgttagcaga attgtcatgc aagggctccc tagctactgg agaatatact
     1381 aagggtactg ttgacattgc gaagagcgac aaagattttg ttatcggctt tattgctcaa
     1441 agagacatgg gtggaagaga tgaaggttac gattggttga ttatgacacc cggtgtgggt
     1501 ttagatgaca agggagacgc attgggtcaa cagtatagaa ccgtggatga tgtggtctct
     1561 acaggatctg acattattat tgttggaaga ggactatttg caaagggaag ggatgctaag
     1621 gtagagggtg aacgttacag aaaagcaggc tgggaagcat atttgagaag atgcggccag
     1681 caaaactaaa aaactgtatt ataagtaaat gcatgtatac taaactcaca aattagagct
     1741 tcaatttaat tatatcagtt attacccata acttcgtata gcatacatta tacgaagtta
     1801 tcccgggtac cgagctcgaa ttcatcacgt gctataaaaa taattataat ttaaattttt
     1861 taatataaat atataaatta aaaatagaaa gtaaaaaaag aaattaaaga aaaaatagtt
     1921 tttgttttcc gaagatgtaa aagactctag ggggatcgcc aacaaatact accttttatc
     1981 ttgctcttcc tgctctcagg tattaatgcc gaattgtttc atcttgtctg tgtagaagac
     2041 cacacacgaa aatcctgtga ttttacattt tacttatcgt taatcgaatg tatatctatt
     2101 taatctgctt ttcttgtcta ataaatatat atgtaaagta cgctttttgt tgaaattttt
     2161 taaacctttg tttatttttt tttcttcatt ccgtaactct tctaccttct ttatttactt
     2221 tctaaaatcc aaatacaaaa cataaaaata aataaacaca gagtaaattc ccaaattatt
     2281 ccatcattaa aagatacgag gcgcgtgtaa gttacgaatt cgtaatcatg gtcatagctg
     2341 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     2401 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     2461 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     2521 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     2581 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     2641 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     2701 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     2761 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     2821 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     2881 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     2941 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     3001 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     3061 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     3121 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aaggacagta
     3181 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     3241 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     3301 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     3361 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     3421 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     3481 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     3541 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     3601 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     3661 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     3721 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     3781 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     3841 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     3901 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     3961 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     4021 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     4081 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     4141 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     4201 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     4261 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     4321 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     4381 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     4441 caaatagggg ttccgcgcac atttccccga aaagtgccac ctgacgtcta agaaaccatt
     4501 attatcatga cattaaccta taaaaatagg cgtatcacga ggccctttcg tctcgcgcgt
     4561 ttcggtgatg acggtgaaaa cctctgacac atgcagctcc cggagacggt cacagcttgt
     4621 ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg cgtcagcggg tgttggcggg
     4681 tgtcggggct ggcttaacta tgcggcatca gagcagattg tactgagagt gcac
//
