LOCUS       sequence-64883-e        5011 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 5011)
  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 5011)
  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: pXP742
FEATURES             Location/Qualifiers
     source          1..5011
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      602..849
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(618..636)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     protein_bind    complement(895..928)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        946..1162
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     CDS             1163..1966
                     /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(2045..2078)
                     /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(2608..2624)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2608..2624)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2621..2643)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2632..2648
                     /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(2656..2662,2663..2680,2681..2686))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2701..2722
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2839..2856)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3010..3598)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3090..3109)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3769..4560,4561..4629))
                     /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     4392..4411
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4630..4734)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     4802..4820
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(4858..4880)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     4980..4999
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 catatgattt agcggccgca aaacgtaggg gcaaacaaac ggaaaaatcg tttctcaaat
       61 tttctgatgc caagaactct aaccagtctt atctaaaaat tgccttatga tccgtctctc
      121 cggttacagc ctgtgtaact gattaatcct gcctttctaa tcaccattct aatgttttaa
      181 ttaagggatt ttgtcttcat taacggcttt cgctcataaa aatgttatga cgttttgccc
      241 gcaggcggga aaccatccac ttcacgagac tgatctcctc tgccggaaca ccgggcatct
      301 ccaacttata agttggagaa ataagagaat ttcagattga gagaatgaaa aaaaaaaaaa
      361 aaaaaaaggc agaggagagc atagaaatgg ggttcacttt ttggtaaagc tatagcatgc
      421 ctatcacata taaatagagt gccagtagcg acttttttca cactcgaaat actcttacta
      481 ctgctctctt gttgttttta tcacttcttg tttcttcttg gtaaatagaa tatcaagcta
      541 caaaaagcat acaatcaact atcaactatt aactatatcg taatacacaa ctagtctcga
      601 gtcatgtaat tagttatgtc acgcttacat tcacgccctc cccccacatc cgctctaacc
      661 gaaaaggaag gagttagaca acctgaagtc taggtcccta tttatttttt tatagttatg
      721 ttagtattaa gaacgttatt tatatttcaa atttttcttt tttttctgta cagacgcgtg
      781 tacgcatgta acattatact gaaaaccttg cttgagaagg ttttgggacg ctcgaaggct
      841 ttaatttgcg gccaagcttg catgcctgca ggtcgactct agaggatccc cgggataact
      901 tcgtatagca tacattatac gaagttatca gggtccataa agcttttcaa ttcatctttt
      961 ttttttttgt tctttttttt gattccggtt tctttgaaat ttttttgatt cggtaatctc
     1021 cgagcagaag gaagaacgaa ggaaggagca cagacttaga ttggtatata tacgcatatg
     1081 tggtgttgaa gaaacatgaa attgcccagt attcttaacc caactgcaca gaacaaaaac
     1141 ctgcaggaaa cgaagataaa tcatgtcgaa agctacatat aaggaacgtg ctgctactca
     1201 tcctagtcct gttgctgcca agctatttaa tatcatgcac gaaaagcaaa caaacttgtg
     1261 tgcttcattg gatgttcgta ccaccaagga attactggag ttagttgaag cattaggtcc
     1321 caaaatttgt ttactaaaaa cacatgtgga tatcttgact gatttttcca tggagggcac
     1381 agttaagccg ctaaaggcat tatccgccaa gtacaatttt ttactcttcg aagacagaaa
     1441 atttgctgac attggtaata cagtcaaatt gcagtactct gcgggtgtat acagaatagc
     1501 agaatgggca gacattacga atgcacacgg tgtggtgggc ccaggtattg ttagcggttt
     1561 gaagcaggcg gcggaagaag taacaaagga acctagaggc cttttgatgt tagcagaatt
     1621 gtcatgcaag ggctccctag ctactggaga atatactaag ggtactgttg acattgcgaa
     1681 gagcgacaaa gattttgtta tcggctttat tgctcaaaga gacatgggtg gaagagatga
     1741 aggttacgat tggttgatta tgacacccgg tgtgggttta gatgacaagg gagacgcatt
     1801 gggtcaacag tatagaaccg tggatgatgt ggtctctaca ggatctgaca ttattattgt
     1861 tggaagagga ctatttgcaa agggaaggga tgctaaggta gagggtgaac gttacagaaa
     1921 agcaggctgg gaagcatatt tgagaagatg cggccagcaa aactaaaaaa ctgtattata
     1981 agtaaatgca tgtatactaa actcacaaat tagagcttca atttaattat atcagttatt
     2041 acccataact tcgtatagca tacattatac gaagttatcc cgggtaccga gctcgaattc
     2101 atcacgtgct ataaaaataa ttataattta aattttttaa tataaatata taaattaaaa
     2161 atagaaagta aaaaaagaaa ttaaagaaaa aatagttttt gttttccgaa gatgtaaaag
     2221 actctagggg gatcgccaac aaatactacc ttttatcttg ctcttcctgc tctcaggtat
     2281 taatgccgaa ttgtttcatc ttgtctgtgt agaagaccac acacgaaaat cctgtgattt
     2341 tacattttac ttatcgttaa tcgaatgtat atctatttaa tctgcttttc ttgtctaata
     2401 aatatatatg taaagtacgc tttttgttga aattttttaa acctttgttt attttttttt
     2461 cttcattccg taactcttct accttcttta tttactttct aaaatccaaa tacaaaacat
     2521 aaaaataaat aaacacagag taaattccca aattattcca tcattaaaag atacgaggcg
     2581 cgtgtaagtt acgaattcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     2641 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     2701 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     2761 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     2821 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     2881 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     2941 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     3001 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     3061 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     3121 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     3181 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     3241 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     3301 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     3361 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     3421 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
     3481 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     3541 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     3601 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     3661 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     3721 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     3781 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     3841 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     3901 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     3961 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     4021 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     4081 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     4141 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     4201 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     4261 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     4321 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     4381 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     4441 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     4501 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     4561 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     4621 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     4681 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     4741 tccccgaaaa gtgccacctg acgtctaaga aaccattatt atcatgacat taacctataa
     4801 aaataggcgt atcacgaggc cctttcgtct cgcgcgtttc ggtgatgacg gtgaaaacct
     4861 ctgacacatg cagctcccgg agacggtcac agcttgtctg taagcggatg ccgggagcag
     4921 acaagcccgt cagggcgcgt cagcgggtgt tggcgggtgt cggggctggc ttaactatgc
     4981 ggcatcagag cagattgtac tgagagtgca c
//
