LOCUS       sequence-336814-        5848 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  To amplify transcription template containing LacI operators .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5848)
  AUTHORS   Xu W, Yan Y, Artsimovitch I, Dunlap D, Finzi L
  TITLE     Positive supercoiling favors transcription elongation through lac 
            repressor-mediated DNA loops.
  JOURNAL   Nucleic Acids Res. 2022 Mar 21;50(5):2826-2835. doi: 
            10.1093/nar/gkac093.
  PUBMED    35188572
REFERENCE   2  (bases 1 to 5848)
  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"; journalName: 
            "Nucleic Acids Res"; date: "2022-03-21- 21"; volume: "50"; issue: 
            "5"; pages: "2826-2835"
COMMENT     Sequence Label: pDM_N1_400
FEATURES             Location/Qualifiers
     source          1..5848
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             39..1229
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(391..410)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     CDS             1868..2059
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     complement(2053..2075)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     protein_bind    2993..3009
                     /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     complement(3803..3820)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3974..4562)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4054..4073)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4733..5524,5525..5593))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     5356..5375
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5594..5698)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5766..5784
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(5811..5816,5817..5833,5834..5839)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
ORIGIN
        1 tatcacagtt aaattgctaa cgcagtcagg caccgtgtat gaaatctaac aatgcgctca
       61 tcgtcatcct cggcaccgtc accctggatg ctgtaggcat aggcttggtt atgccggtac
      121 tgccgggcct cttgcgggat atcgtccatt ccgacagcat cgccagtcac tatggcgtgc
      181 tgctagcgct atatgcgttg atgcaatttc tatgcgcacc cgttctcgga gcactgtccg
      241 accgctttgg ccgccgccca gtcctgctcg cttcgctact tggagccact atcgactacg
      301 cgatcatggc gaccacaccc gtcctgtgga tcctctacgc cggacgcatc gtggccggca
      361 tcaccggcgc cacaggtgcg gttgctggcg cctatatcgc cgacatcacc gatggggaag
      421 atcgggctcg ccacttcggg ctcatgagcg cttgtttcgg cgtgggtatg gtggcaggcc
      481 ccgtggccgg gggactgttg ggcgccatct ccttgcatgc accattcctt gcggcggcgg
      541 tgctcaacgg cctcaaccta ctactgggct gcttcctaat gcaggagtcg cataagggag
      601 agcgtcgacc gatgcccttg agagccttca acccagtcag ctccttccgg tgggcgcggg
      661 gcatgactat cgtcgccgca cttatgactg tcttctttat catgcaactc gtaggacagg
      721 tgccggcagc gctctgggtc attttcggcg aggaccgctt tcgctggagc gcgacgatga
      781 tcggcctgtc gcttgcggta ttcggaatct tgcacgccct cgctcaagcc ttcgtcactg
      841 gtcccgccac caaacgtttc ggcgagaagc aggccattat cgccggcatg gcggccgacg
      901 cgctgggcta cgtcttgctg gcgttcgcga cgcgaggctg gatggccttc cccattatga
      961 ttcttctcgc ttccggcggc atcgggatgc ccgcgttgca ggccatgctg tccaggcagg
     1021 tagatgacga ccatcaggga cagcttcaag gatcgctcgc ggctcttacc agcctaactt
     1081 cgatcattgg accgctgatc gtcacggcga tttatgccgc ctcggcgagc acatggaacg
     1141 ggttggcatg gattgtaggc gccgccctat accttgtctg cctccccgcg ttgcgtcgcg
     1201 gtgcatggag ccgggccacc tcgacctgaa tggaagccgg cggcacctcg ctaacggatt
     1261 caccactcca agaattggag ccaatcaatt cttgcggaga actgtgaatg cgcaaaccaa
     1321 cccttggcag aacgtatcca tcgcgtccgc catctccagc agccgcacgc ggcgcatctc
     1381 gggcagcgtt gggtcctggc cacgggtgcg catgatcgtg ctcctgtcgt tgaggacccg
     1441 gctaggctgg cggggttgcc ttactggtta gcagaatgaa tcaccgatac gcgagcgaac
     1501 gtgaagcgac tgctgctgca aaacgtctgc gacctgagca acaacatgaa tggtcttcgg
     1561 tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca ttatgttccg
     1621 gatctgcatc gcaggatgct gctggctacc ctgtggaaca cctacatctg tattaacgaa
     1681 gcgctggcat tgaccctgag tgatttttct ctggtcccgc cgcatccata ccgccagttg
     1741 tttaccctca caacgttcca gtaaccgggc atgttcatca tcagtaaccc gtatcgtgag
     1801 catcctctct cgtttcatcg gtatcattac ccccatgaac agaaatcccc cttacacgga
     1861 ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg gcccgcttta tcagaagcca
     1921 gacattaacg cttctggaga aactcaacga gctggacgcg gatgaacagg cagacatctg
     1981 tgaatcgctt cacgaccacg ctgatgagct ttaccgcagc tgcctcgcgc gtttcggtga
     2041 tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt gtctgtaagc
     2101 ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggca ggtgtcgggg
     2161 cgcagccatg acccagtcac cccatggtgc agtatgaagg cggcggagcc gacaccacgg
     2221 ccaccgatat tatttgcccg atgtacgcgc gcgtggatga agaccagccc ttcccggctt
     2281 tatcaaaaag agtattgact taaagtctaa cctataggat acttacagcg atggagaggt
     2341 gtagtggtaa ccagaagata atatggcttt cgctacctgg agagacgcgc ccgctgatcc
     2401 tttgcgaata cgcccacgcg atgggtaaca gtcttggcgg tttcgctaaa tactggcagg
     2461 cgtttcgtca gtatccccgt ttacagggcg gcttcgtctg ggactgggtg gatcagtcgc
     2521 tgattaaata tgatgaaaac ggcaacccgt ggtaccggct tacggcggtg atttttgcga
     2581 tacgccgaac gtcgatcgag ctcaacgcaa ttatcgccag ttctgtatga acggtctggt
     2641 ctttgccgac cgcacgtcgc atccagcgct gacggaagca aaacaccagc agcagttttt
     2701 cagttccgtt tatccgggca aaccatcgaa gtgaccagcg aatacctgtt ccgtcatagc
     2761 gataacgagc tcctgcactg gatggtggcg ctggatggta agccgctggc aagcggtgaa
     2821 gtgcctctgg atgtcgctcc acaaggtaaa cagttgattg aactgtctga actaccgcag
     2881 ccggagagcg ccgggcaact ctggctcaca gtacgcgtag tgcaaccgaa cgcgaccgca
     2941 tggtcagaac tagatgtaca gaagtcagat tagcagtagc gtctggcgga aattgtgagc
     3001 ggataacaat taaacctcag tgtgacgctc cccgccgcgt cccacgccat cccgcatctg
     3061 accaccagcg aaatggattt ttgcactcga gctgggtaat aagcgttggc cccgacaact
     3121 ccatctgctt cttccgtgcg agatgtggat tggcgataaa aaacaactgc tgacgccgct
     3181 gcgcgatcag ttcacccgtg caccgctgga taacgacatt ggcgtaagtg aagcgacccg
     3241 cattgaccct aacgcctggg tcgaacactg gaaggcggcg ggccattacc aggccgaagc
     3301 agcgcgccgc ttgcgatttc gaccaacagc tgctgatgcg gtgctgatta cgaccgctca
     3361 cgcgtggcag catcagggga aaaccttatt tatcagccgg aagcaccgtg taacttccag
     3421 gacatctatg atggcgatta ccgttgatgt tgaagtggcg agcgatacac cgcatccggc
     3481 gcggatttct gaaaagatgt cgtgctactc catactcggg gtcctgtatg ggctcggatt
     3541 agcggccgca agaaaactat cccgaccgcc ttactgccgc ctgttttgac cgctgggatc
     3601 tgctgtaaca gagcattagc gcaaggtgat ttttgtcttc ttgcgctaat tttttccact
     3661 ggatctacag aagcgcgatt gtcgggcaca ggcttaacta tgcggcatca gagcagacag
     3721 agtacgacga tttgcgatgg aacttggaga taccgcacag atgcgtaagg agaaaatacc
     3781 gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc gttcggctgc
     3841 ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa tcaggggata
     3901 acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg
     3961 cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct
     4021 caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa
     4081 gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc
     4141 tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt
     4201 aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg
     4261 ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg
     4321 cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct
     4381 tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc tgcgctctgc
     4441 tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg
     4501 ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc
     4561 aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt
     4621 aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa
     4681 aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat
     4741 gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct
     4801 gactccccgt cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg
     4861 caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata aaccagccag
     4921 ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta
     4981 attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg
     5041 ccattgctgc aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg
     5101 gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct
     5161 ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta
     5221 tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg
     5281 gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc
     5341 cggcgtcaac acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg
     5401 gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga
     5461 tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg
     5521 ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat
     5581 gttgaatact catactcttc ctttttcaat attattgaag catttatcag ggttattgtc
     5641 tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca
     5701 catttccccg aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct
     5761 ataaaaatag gcgtatcacg aggccctttc gtcttcaaga attctcatgt ttgacagctt
     5821 atcatcgata agctttaatg cggtagtt
//
