LOCUS       sequence-42843-e        8063 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 8063)
  AUTHORS   Larson DR, Zenklusen D, Wu B, Chao JA, Singer RH
  TITLE     Real-time observation of transcription initiation and elongation on 
            an endogenous yeast gene.
  JOURNAL   Science. 2011 Apr 22;332(6028):475-8.
  PUBMED    21512033
REFERENCE   2  (bases 1 to 8063)
  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"; journalName: "Science."; 
            date: "2011-04-22"; volume: "332(6028)"; pages: "475-8"
COMMENT     Sequence Label: pDZ207 (loxP KAN loxP 24xPP7SL MDN1)
FEATURES             Location/Qualifiers
     source          1..8063
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        242..260
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     protein_bind    712..745
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     gene            800..2156
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin 
                     resistance (Wach et al., 1994)"
     promoter        800..1143
                     /label=TEF promoter
                     /note="Ashbya gossypii TEF promoter"
     CDS             1144..1953
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP
                     DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA
                     FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD
                     FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD
                     RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     complement(1211..1230)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     terminator      1959..2156
                     /label=TEF terminator
                     /note="Ashbya gossypii TEF terminator"
     protein_bind    2219..2252
                     /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(4437..4456)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4438..4456)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4463..4480)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4463..4479)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4472..4494)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             4617..4919
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     4798..4817
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             5268..6062
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             5268..6062
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5322..5341)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     5932..5951
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     CDS             complement(6608..6986)
                     /label=ISS
                     /note="immunostimulatory sequence from the AmpR gene; 
                     contains unmethylated CpG dinucleotides in the context of 
                     5'-AACGTT-3' (Sato et al., 1996)"
     primer_bind     6933..6952
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7294..7882
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7783..7802
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8036..8053
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctca
      241 gaattaaccc tcactaaagg gactagtcct gcaggtttaa acgaattcgc ccttatatat
      301 ttaattaaag atcaaccggt ctcaacaaac ttcccatact aaagctttct cttgatatat
      361 agtcaaactg aatgtatact gaaacacata ttcaaggcat tgattttaaa tgcacttgaa
      421 ccgttattcc atttcttttt ctactcatca tttattgtcg ccctttacga ggtatttttt
      481 ctttttcgag tgaaaaattt taaagaagcg atgagctaag gtctatttag ctttatgaaa
      541 agaaattata tcacttactg ctttacttta gacattcctc tccaaagtga ttattttaaa
      601 gattcaaagt acctctgagt taatttgaca tcgaattcta ctcttattat tttaaggtac
      661 ctctacttta cgcggtcaca tctcgagggc tgcaggtcga caacccttaa tataacttcg
      721 tataatgtat gctatacgaa gttattaggt ctagagatct gtttagcttg cctcgtcccc
      781 gccgggtcac ccggccagcg acatggaggc ccagaatacc ctccttgaca gtcttgacgt
      841 gcgcagctca ggggcatgat gtgactgtcg cccgtacatt tagcccatac atccccatgt
      901 ataatcattt gcatccatac attttgatgg ccgcacggcg cgaagcaaaa attacggctc
      961 ctcgctgcag acctgcgagc agggaaacgc tcccctcaca gacgcgttga attgtcccca
     1021 cgccgcgccc ctgtagagaa atataaaagg ttaggatttg ccactgaggt tcttctttca
     1081 tatacttcct tttaaaatct tgctaggata cagttctcac atcacatccg aacataaaca
     1141 accatgggta aggaaaagac tcacgtttcg aggccgcgat taaattccaa catggatgct
     1201 gatttatatg ggtataaatg ggctcgcgat aatgtcgggc aatcaggtgc gacaatctat
     1261 cgattgtatg ggaagcccga tgcgccagag ttgtttctga aacatggcaa aggtagcgtt
     1321 gccaatgatg ttacagatga gatggtcaga ctaaactggc tgacggaatt tatgcctctt
     1381 ccgaccatca agcattttat ccgtactcct gatgatgcat ggttactcac cactgcgatc
     1441 cccggcaaaa cagcattcca ggtattagaa gaatatcctg attcaggtga aaatattgtt
     1501 gatgcgctgg cagtgttcct gcgccggttg cattcgattc ctgtttgtaa ttgtcctttt
     1561 aacagcgatc gcgtatttcg tctcgctcag gcgcaatcac gaatgaataa cggtttggtt
     1621 gatgcgagtg attttgatga cgagcgtaat ggctggcctg ttgaacaagt ctggaaagaa
     1681 atgcataagc ttttgccatt ctcaccggat tcagtcgtca ctcatggtga tttctcactt
     1741 gataacctta tttttgacga ggggaaatta ataggttgta ttgatgttgg acgagtcgga
     1801 atcgcagacc gataccagga tcttgccatc ctatggaact gcctcggtga gttttctcct
     1861 tcattacaga aacggctttt tcaaaaatat ggtattgata atcctgatat gaataaattg
     1921 cagtttcatt tgatgctcga tgagtttttc taatcagtac tgacaataaa aagattcttg
     1981 ttttcaagaa cttgtcattt gtatagtttt tttatattgt agttgttcta ttttaatcaa
     2041 atgttagcgt gatttatatt ttttttcgcc tcgacatcat ctgcccagat gcgaagttaa
     2101 gtgcgcagaa agtaatatca tgcgtcaatc gtatgtgaat gctggtcgct atactgctgt
     2161 cgattcgata ctaacgccgc catccagtgt cgaaaacgag ctctcgagaa cccttaatat
     2221 aacttcgtat aatgtatgct atacgaagtt attaggtgat atcagatcca ctagtgggga
     2281 tcctaaggta cctaattgcc tagaaaggag cagacgatat ggcgtcgctc cctgcaggtc
     2341 gactctagaa accagcagag catatgggct cgctggctgc agtattcccg ggttcattag
     2401 atcctaaggt acctaattgc ctagaaagga gcagacgata tggcgtcgct ccctgcaggt
     2461 cgactctaga aaccagcaga gcatatgggc tcgctggctg cagtattccc gggttcatta
     2521 gatcctaagg tacctaattg cctagaaagg agcagacgat atggcgtcgc tccctgcagg
     2581 tcgactctag aaaccagcag agcatatggg ctcgctggct gcagtattcc cgggttcatt
     2641 agatcctaag gtacctaatt gcctagaaag gagcagacga tatggcgtcg ctccctgcag
     2701 gtcgactcta gaaaccagca gagcatatgg gctcgctggc tgcagtattc ccgggttcat
     2761 tagatcctaa ggtacctaat tgcctagaaa ggagcagacg atatggcgtc gctccctgca
     2821 ggtcgactct agaaaccagc agagcatatg ggctcgctgg ctgcagtatt cccgggttca
     2881 ttagatccta aggtacctaa ttgcctagaa aggagcagac gatatggcgt cgctccctgc
     2941 aggtcgactc tagaaaccag cagagcatat gggctcgctg gctgcagtat tcccgggttc
     3001 attagatcct aaggtaccta attgcctaga aaggagcaga cgatatggcg tcgctccctg
     3061 caggtcgact ctagaaacca gcagagcata tgggctcgct ggctgcagta ttcccgggtt
     3121 cattagatcc taaggtacct aattgcctag aaaggagcag acgatatggc gtcgctccct
     3181 gcaggtcgac tctagaaacc agcagagcat atgggctcgc tggctgcagt attcccgggt
     3241 tcattagatc ctaaggtacc taattgccta gaaaggagca gacgatatgg cgtcgctccc
     3301 tgcaggtcga ctctagaaac cagcagagca tatgggctcg ctggctgcag tattcccggg
     3361 ttcattagat cctaaggtac ctaattgcct agaaaggagc agacgatatg gcgtcgctcc
     3421 ctgcaggtcg actctagaaa ccagcagagc atatgggctc gctggctgca gtattcccgg
     3481 gttcattaga tcctaaggta cctaattgcc tagaaaggag cagacgatat ggcgtcgctc
     3541 cctgcaggtc gactctagaa accagcagag catatgggct cgctggctgc agtattcccg
     3601 ggttcattag atcctaaggt acctaattgc ctagaaagga gcagacgata tggcgtcgct
     3661 ccctgcaggt cgactctaga aaccagcaga gcatatgggc tcgctggctg cagtattccc
     3721 gggttcatta gatccatgtc ccaggataga attttgttag atttagacgt agttaaccaa
     3781 aggttaattc tcttcaattc tgcttttccc tctgatgcta ttgaagcgcc cttccacttt
     3841 agcaacaagg aatcaacttc tgagaattta gacaatttag caggaactat actacatagc
     3901 cgttccatca ctggtcatgt tttcctttac aaacatattt ttttagagat agtggcacgt
     3961 tggataaagg actccaaaaa aaaagattac gttcttgtaa ttgaaaagtt ggcaagtata
     4021 attacaatat ttcccgttgc aatgccatta attgaagact atttagataa agagaatgat
     4081 catttcatca ctattcttca aaatccatcc acacaaaagg attccgacat gtttaaaata
     4141 ctattagcat actataggtt actatatcac aataaagaag tatttgcacg cttcattcaa
     4201 cccgacattt tataccaact tgtggattta ctcaccaaag aacaagaaaa tcaggttgtt
     4261 atatttctag cattaaaggt tttatcttta tacttagaca tgggagaaaa aacattaaac
     4321 gatatgctcg acacttatat aaaatcacgc gattcattgt tgggccattt tgagggcgat
     4381 tctggaattg attacagctt cctaagggcg aattcgcggc cgctaaattc aattcgccct
     4441 atagtgagtc gtattacaat tcactggccg tcgttttaca acgtcgtgac tgggaaaacc
     4501 ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc tggcgtaata
     4561 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctatac gtacggcagt
     4621 ttaaggttta cacctataaa agagagagcc gttatcgtct gtttgtggat gtacagagtg
     4681 atattattga cacgccgggg cgacggatgg tgatccccct ggccagtgca cgtctgctgt
     4741 cagataaagt ctcccgtgaa ctttacccgg tggtgcatat cggggatgaa agctggcgca
     4801 tgatgaccac cgatatggcc agtgtgccgg tctccgttat cggggaagaa gtggctgatc
     4861 tcagccaccg cgaaaatgac atcaaaaacg ccattaacct gatgttctgg ggaatataaa
     4921 tgtcaggcat gagattatca aaaaggatct tcacctagat ccttttcacg tagaaagcca
     4981 gtccgcagaa acggtgctga ccccggatga atgtcagcta ctgggctatc tggacaaggg
     5041 aaaacgcaag cgcaaagaga aagcaggtag cttgcagtgg gcttacatgg cgatagctag
     5101 actgggcggt tttatggaca gcaagcgaac cggaattgcc agctggggcg ccctctggta
     5161 aggttgggaa gccctgcaaa gtaaactgga tggctttctt gccgccaagg atctgatggc
     5221 gcaggggatc aagctctgat caagagacag gatgaggatc gtttcgcatg attgaacaag
     5281 atggattgca cgcaggttct ccggccgctt gggtggagag gctattcggc tatgactggg
     5341 cacaacagac aatcggctgc tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc
     5401 cggttctttt tgtcaagacc gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag
     5461 cgcggctatc gtggctggcc acgacgggcg ttccttgcgc agctgtgctc gacgttgtca
     5521 ctgaagcggg aagggactgg ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat
     5581 ctcaccttgc tcctgccgag aaagtatcca tcatggctga tgcaatgcgg cggctgcata
     5641 cgcttgatcc ggctacctgc ccattcgacc accaagcgaa acatcgcatc gagcgagcac
     5701 gtactcggat ggaagccggt cttgtcgatc aggatgatct ggacgaagag catcaggggc
     5761 tcgcgccagc cgaactgttc gccaggctca aggcgagcat gcccgacggc gaggatctcg
     5821 tcgtgaccca tggcgatgcc tgcttgccga atatcatggt ggaaaatggc cgcttttctg
     5881 gattcatcga ctgtggccgg ctgggtgtgg cggaccgcta tcaggacata gcgttggcta
     5941 cccgtgatat tgctgaagag cttggcggcg aatgggctga ccgcttcctc gtgctttacg
     6001 gtatcgccgc tcccgattcg cagcgcatcg ccttctatcg ccttcttgac gagttcttct
     6061 gaattattaa cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta
     6121 tttcacaccg catcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     6181 tttttctaaa tacattcaaa tatgtatccg ctcatgagat tatcaaaaag gatcttcacc
     6241 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     6301 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     6361 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     6421 ccatctggcc ccagtgctgc aatgataccg cgagaccacg ctcaccggct ccagatttat
     6481 cagcaataaa ccagccagcc ggaaggccga gcgcagaagt ggtcctgcaa ctttatccgc
     6541 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     6601 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     6661 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     6721 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     6781 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     6841 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     6901 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     6961 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     7021 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     7081 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     7141 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     7201 ttatcagggt tattgtctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc
     7261 gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat
     7321 ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga
     7381 gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt
     7441 tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata
     7501 cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac
     7561 cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg
     7621 ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg
     7681 tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag
     7741 cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct
     7801 ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc
     7861 aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt
     7921 ttgctggcct tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg
     7981 tattaccgcc tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga
     8041 gtcagtgagc gaggaagcgg aag
//
