LOCUS       sequence-330577-        5849 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 5849)
  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 5849)
  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: pRS_1N_400
FEATURES             Location/Qualifiers
     source          1..5849
                     /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    2594..2610
                     /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     3704..3723
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(3804..3821)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3975..4563)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4055..4074)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4734..5525,5526..5594))
                     /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     5357..5376
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5595..5699)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     5767..5785
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        join(5812..5817,5818..5834,5835..5840)
                     /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 aacatatcca 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 acaccagccc ttcccggctt
     2281 tatcaaaaag agtattgact taaagtctaa cctataggat acttacagcg atggagaggt
     2341 gtagtggtaa ccagaagata agatggcttt cgctacctgg agagacgcgc ccgctgatcc
     2401 tttgcgaata cgcccacgcg atgggtaaca gtcttggcgg tttcgctaaa tactggcagg
     2461 cgtttcgtca gtatccccgt ttacagggcg gcttcgtctg ggactgggtg gatcagtcgc
     2521 tgattaaata tgatgaaaac ggcaacccgt ggtaccggct tacggcggtg attttggcga
     2581 tacgccgaac gaattgtgag cggataacaa ttatcgccag ttctgtatga acggtctggt
     2641 ctttgccgac cgcacgccgc atccagcgct gacggaagca aaacaccagc agcagttttt
     2701 ccagttccgt ttatccgggc aaaccatcga agtgaccagc gaatacctgt tccgtcatag
     2761 cgataacgag ctcctgcact ggatggtggc gctggatggt aagccgctgg caagcggtga
     2821 agtgcctctg gatgtcgctc cacaaggtaa acagttgatt gaactgcctg aactaccgca
     2881 gccggagagc gccgggcaac tctggctcac agtacgcgta gtgcaaccga acgcgaccgc
     2941 atggtcagaa gccgggcaca tcagcgcctg gcagcagtag cgtctggcgg atcgatcgag
     3001 ctcaacgcaa ttaaacctca gtgtgacgct ccccgccgcg tcccacgcca tcccgcatct
     3061 gaccaccagc gaaatggatt tttgcactcg agctgggtaa taagcgttgg caatttaacc
     3121 gccagtcagg ctttctttca cagatgtgga ttggcgataa aaaacaactg ctgacgccgc
     3181 tgcgcgatca gttcacccgt gcaccgctgg ataacgacat tggcgtaagt gaagcgaccc
     3241 gcattgaccc taacgcctgg gtcgaacgct ggaaggcggc gggccattac caggccgaag
     3301 cagcgttgtt gcagtgcacg gcagatacac ttgctgatgc ggtgctgatt acgaccgctc
     3361 acgcgtggca gcatcagggg aaaaccttat ttatcagccg gaaaacctac cggattgatg
     3421 gtagtggtca aatggcgatt accgttgatg ttgaagtggc gagcgataca ccgcatccgg
     3481 cgcggattgg cctgaactgc cagctggcgc aggtagcaga gcgggtaaac tggctcggat
     3541 tagcggccgc aagaaaacta tcccgaccgc cttactgccg cctgttttga ccgctgggat
     3601 ctgctgtaac agagcattag cgcaaggtga tttttgtctt cttgcgctaa ttttttccat
     3661 tgtctagagt agcgatagcg gagtgtatac tggcttaact atgcggcatc agagcagatt
     3721 gtactgagag tgcaccatat gcggtgtgaa ataccgcaca gatgcgtaag gagaaaatac
     3781 cgcatcaggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     3841 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     3901 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     3961 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4021 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4081 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4141 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4201 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     4261 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4321 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4381 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     4441 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     4501 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     4561 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     4621 taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct tttaaattaa
     4681 aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga cagttaccaa
     4741 tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc catagttgcc
     4801 tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg ccccagtgct
     4861 gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat aaaccagcca
     4921 gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat ccagtctatt
     4981 aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg caacgttgtt
     5041 gccattgctg caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc
     5101 ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa agcggttagc
     5161 tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc actcatggtt
     5221 atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt ttctgtgact
     5281 ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc
     5341 ccggcgtcaa cacgggataa taccgcgcca catagcagaa ctttaaaagt gctcatcatt
     5401 ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag atccagttcg
     5461 atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac cagcgtttct
     5521 gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa
     5581 tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca gggttattgt
     5641 ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc
     5701 acatttcccc gaaaagtgcc acctgacgtc taagaaacca ttattatcat gacattaacc
     5761 tataaaaata ggcgtatcac gaggcccttt cgtcttcaag aattctcatg tttgacagct
     5821 tatcatcgat aagctttaat gcggtagtt
//
