LOCUS       sequence-65292-e        5735 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 5735)
  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 5735)
  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: pXP822
FEATURES             Location/Qualifiers
     source          1..5735
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        8..450
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    8..125
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     421..444
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     terminator      471..718
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(487..505)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    complement(764..797)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        815..1031
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             1032..1835
                     /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(1914..1947)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      1970..3312
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    complement(2933..2980)
                     /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     complement(3332..3348)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3332..3348)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3345..3367)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3356..3372
                     /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(3380..3386,3387..3404,3405..3410))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3425..3446
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3563..3580)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3734..4322)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3814..3833)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4493..5284,5285..5353))
                     /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     5116..5135
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5354..5458)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5526..5544
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(5582..5604)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     5704..5723
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 catatgacgg attagaagcc gccgagcggg tgacagccct ccgaaggaag actctcctcc
       61 gtgcgtcctc gtcctcaccg gtcgcgttcc tgaaacgcag atgtgcctcg cgccgcactg
      121 ctccgaacaa taaagattct acaatactag cttttatggt tatgaagagg aaaaattggc
      181 agtaacctgg ccccacaaac cttcaaatga acgaatcaaa ttaacaacca taggatgata
      241 atgcgattag ttttttagcc ttatttctgg ggtaattaat cagcgaagcg atgatttttg
      301 atctattaac agatatataa atgcaaaaac tgcattaacc actttaacta atactttcaa
      361 cattttcggt ttgtattact tcttattcaa atgtaataaa agtatcaaca aaaaattgtt
      421 aatatacctc tatactttaa cgtcaaggag aaaaaaccac tagtctcgag tcatgtaatt
      481 agttatgtca cgcttacatt cacgccctcc ccccacatcc gctctaaccg aaaaggaagg
      541 agttagacaa cctgaagtct aggtccctat ttattttttt atagttatgt tagtattaag
      601 aacgttattt atatttcaaa tttttctttt ttttctgtac agacgcgtgt acgcatgtaa
      661 cattatactg aaaaccttgc ttgagaaggt tttgggacgc tcgaaggctt taatttgcgg
      721 ccaagcttgc atgcctgcag gtcgactcta gaggatcccc gggataactt cgtatagcat
      781 acattatacg aagttatcag ggtccataaa gcttttcaat tcatcttttt tttttttgtt
      841 cttttttttg attccggttt ctttgaaatt tttttgattc ggtaatctcc gagcagaagg
      901 aagaacgaag gaaggagcac agacttagat tggtatatat acgcatatgt ggtgttgaag
      961 aaacatgaaa ttgcccagta ttcttaaccc aactgcacag aacaaaaacc tgcaggaaac
     1021 gaagataaat catgtcgaaa gctacatata aggaacgtgc tgctactcat cctagtcctg
     1081 ttgctgccaa gctatttaat atcatgcacg aaaagcaaac aaacttgtgt gcttcattgg
     1141 atgttcgtac caccaaggaa ttactggagt tagttgaagc attaggtccc aaaatttgtt
     1201 tactaaaaac acatgtggat atcttgactg atttttccat ggagggcaca gttaagccgc
     1261 taaaggcatt atccgccaag tacaattttt tactcttcga agacagaaaa tttgctgaca
     1321 ttggtaatac agtcaaattg cagtactctg cgggtgtata cagaatagca gaatgggcag
     1381 acattacgaa tgcacacggt gtggtgggcc caggtattgt tagcggtttg aagcaggcgg
     1441 cggaagaagt aacaaaggaa cctagaggcc ttttgatgtt agcagaattg tcatgcaagg
     1501 gctccctagc tactggagaa tatactaagg gtactgttga cattgcgaag agcgacaaag
     1561 attttgttat cggctttatt gctcaaagag acatgggtgg aagagatgaa ggttacgatt
     1621 ggttgattat gacacccggt gtgggtttag atgacaaggg agacgcattg ggtcaacagt
     1681 atagaaccgt ggatgatgtg gtctctacag gatctgacat tattattgtt ggaagaggac
     1741 tatttgcaaa gggaagggat gctaaggtag agggtgaacg ttacagaaaa gcaggctggg
     1801 aagcatattt gagaagatgc ggccagcaaa actaaaaaac tgtattataa gtaaatgcat
     1861 gtatactaaa ctcacaaatt agagcttcaa tttaattata tcagttatta cccataactt
     1921 cgtatagcat acattatacg aagttatccc gggtaccgag ctcgaattca acgaagcatc
     1981 tgtgcttcat tttgtagaac aaaaatgcaa cgcgagagcg ctaatttttc aaacaaagaa
     2041 tctgagctgc atttttacag aacagaaatg caacgcgaaa gcgctatttt accaacgaag
     2101 aatctgtgct tcatttttgt aaaacaaaaa tgcaacgcga gagcgctaat ttttcaaaca
     2161 aagaatctga gctgcatttt tacagaacag aaatgcaacg cgagagcgct attttaccaa
     2221 caaagaatct atacttcttt tttgttctac aaaaatgcat cccgagagcg ctatttttct
     2281 aacaaagcat cttagattac tttttttctc ctttgtgcgc tctataatgc agtctcttga
     2341 taactttttg cactgtaggt ccgttaaggt tagaagaagg ctactttggt gtctattttc
     2401 tcttccataa aaaaagcctg actccacttc ccgcgtttac tgattactag cgaagctgcg
     2461 ggtgcatttt ttcaagataa aggcatcccc gattatattc tataccgatg tggattgcgc
     2521 atactttgtg aacagaaagt gatagcgttg atgattcttc attggtcaga aaattatgaa
     2581 cggtttcttc tattttgtct ctatatacta cgtataggaa atgtttacat tttcgtattg
     2641 ttttcgattc actctatgaa tagttcttac tacaattttt ttgtctaaag agtaatacta
     2701 gagataaaca taaaaaatgt agaggtcgag tttagatgca agttcaagga gcgaaaggtg
     2761 gatgggtagg ttatataggg atatagcaca gagatatata gcaaagagat acttttgagc
     2821 aatgtttgtg gaagcggtat tcgcaatatt ttagtagctc gttacagtcc ggtgcgtttt
     2881 tggttttttg aaagtgcgtc ttcagagcgc ttttggtttt caaaagcgct ctgaagttcc
     2941 tatactttct agagaatagg aacttcggaa taggaacttc aaagcgtttc cgaaaacgag
     3001 cgcttccgaa aatgcaacgc gagctgcgca catacagctc actgttcacg tcgcacctat
     3061 atctgcgtgt tgcctgtata tatatataca tgagaagaac ggcatagtgc gtgtttatgc
     3121 ttaaatgcgt acttatatgc gtctatttat gtaggatgaa aggtagtcta gtacctcctg
     3181 tgatattatc ccattccatg cggggtatcg tatgcttcct tcagcactac cctttagctg
     3241 ttctatatgc tgccactcct caattggatt agtctcatcc ttcaatgcta tcatttcctt
     3301 tgatattgga tcatacgaat tcgtaatcat ggtcatagct gtttcctgtg tgaaattgtt
     3361 atccgctcac aattccacac aacatacgag ccggaagcat aaagtgtaaa gcctggggtg
     3421 cctaatgagt gagctaactc acattaattg cgttgcgctc actgcccgct ttccagtcgg
     3481 gaaacctgtc gtgccagctg cattaatgaa tcggccaacg cgcggggaga ggcggtttgc
     3541 gtattgggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3601 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3661 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3721 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     3781 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     3841 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     3901 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     3961 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4021 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4081 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4141 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     4201 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4261 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4321 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4381 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4441 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4501 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4561 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4621 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4681 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4741 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     4801 ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     4861 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     4921 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     4981 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5041 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5101 cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5161 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5221 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5281 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5341 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5401 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5461 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     5521 ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa
     5581 acctctgaca catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga
     5641 gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact
     5701 atgcggcatc agagcagatt gtactgagag tgcac
//
