LOCUS       sequence-149510-        4271 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Dual selection plasmid with lac operator, selection for binding with
            spectinomycin, for non-binding with chloramphenicol.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4271)
  AUTHORS   Hoffmann SA, Kruse SM, Arndt KM
  TITLE     Long-range transcriptional interference in E. coli used to construct
            a dual positive selection system for genetic switches.
  JOURNAL   Nucleic Acids Res. 2016 Jun 2;44(10):e95. doi: 10.1093/nar/gkw125. 
            Epub 2016 Feb 29.
  PUBMED    26932362
REFERENCE   2  (bases 1 to 4271)
  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: "Nucleic 
            Acids Res."; date: "2016-06-2"; volume: "44(10):e95. doi"; pages: " 
            10.1093/nar/gkw125. Epub 2016 Feb 29"
COMMENT     Sequence Label: pSAH0016
FEATURES             Location/Qualifiers
     source          1..4271
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(202..1152)
                     /codon_start=1
                     /gene="rep101"
                     /product="RepA protein needed for replication with the 
                     pSC101 origin"
                     /label=Rep101
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYAQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      1200..1422
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     terminator      complement(1914..2008)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     terminator      complement(2164..2193)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     promoter        join(2256..2261,2262..2277,2278..2284)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2290..2309
                     /label=lac operator (symmetric)
                     /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). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     CDS             2335..2994
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2401..2420)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             complement(3001..3792)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVIGK"
     terminator      4146..4177
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
ORIGIN
        1 gttacattgt cgatctgttc atggtgaaca gctttgaatg caccaaaaac tcgtaaaagc
       61 tctgatgtat ctatcttttt tacaccgttt tcatctgtgc atatggacag ttttcccttt
      121 gatatgtaac ggtgaacagt tgttctactt ttgtttgtta gtcttgatgc ttcactgata
      181 gatacaagag ccataagaac ctcagatcct tccgtattta gccagtatgt tctctagtgt
      241 ggttcgttgt ttttgcgtga gccatgagaa cgaaccattg agatcatact tactttgcat
      301 gtcactcaaa aattttgcct caaaactggt gagctgaatt tttgcagtta aagcatcgtg
      361 tagtgttttt cttagtccgt tatgtaggta ggaatctgat gtaatggttg ttggtatttt
      421 gtcaccattc atttttatct ggttgttctc aagttcggtt acgagatcca tttgtctatc
      481 tagttcaact tggaaaatca acgtatcagt cgggcggcct cgcttatcaa ccaccaattt
      541 catattgctg taagtgttta aatctttact tattggtttc aaaacccatt ggttaagcct
      601 tttaaactca tggtagttat tttcaagcat taacatgaac ttaaattcat caaggctaat
      661 ctctatattt gccttgtgag ttttcttttg tgttagttct tttaataacc actcataaat
      721 cctcatagag tatttgtttt caaaagactt aacatgttcc agattatatt ttatgaattt
      781 ttttaactgg aaaagataag gcaatatctc ttcactaaaa actaattcta atttttcgct
      841 tgagaacttg gcatagtttg tccactggaa aatctcaaag cctttaacca aaggattcct
      901 gatttccaca gttctcgtca tcagctctct ggttgcttta gctaatacac cataagcatt
      961 ttccctactg atgttcatca tctgagcgta ttggttataa gtgaacgata ccgtccgttc
     1021 tttccttgta gggttttcaa tcgtggggtt gagtagtgcc acacagcata aaattagctt
     1081 ggtttcatgc tccgttaagt catagcgact aatcgctagt tcatttgctt tgaaaacaac
     1141 taattcagac atacatctca attggtctag gtgattttaa tcactatacc aattgagatg
     1201 ggctagtcaa tgataattac atgtcctttt cctttgagtt gtgggtatct gtaaattctg
     1261 ctagaccttt gctggaaaac ttgtaaattc tgctagaccc tctgtaaatt ccgctagacc
     1321 tttgtgtgtt ttttttgttt atattcaagt ggttataatt tatagaataa agaaagaata
     1381 aaaaaagata aaaagaatag atcccagccc tgtgtataac tcactacttt agtcagttcc
     1441 gcagtattac aaaaggatgt cgcaaacgct gtttgctcct ctacaaaaca gaccttaaaa
     1501 ccctaaaggc ttaagtagca ccctcgcaag ctcgggcaaa tcgctgaata ttccttttgt
     1561 ctccgaccat caggcacctg agtcgctgtc tttttcgtga cattcagttc gctgcgctca
     1621 cggctctggc agtgaatggg ggtaaatggc actacaggcg ccttttatgg attcatgcaa
     1681 ggaaactacc cataatacaa gaaaagcccg tcacgggctt ctcagggcgt tttatggcgg
     1741 gtctgctatg tggtgctatc tgactttttg ctgttcagca gttcctgccc tctgattttc
     1801 cagtctgacc acttcggatt atcccgtgac aggtcattca gactggctaa tgcacccagt
     1861 aaggcagcgg tatcatcaac aggcttaccc gtcttactgt ccctagtgct tggattctca
     1921 ccaataaaaa acgcccggcg gcaaccgagc gttctgaaca aatccagatg gagttctgag
     1981 gtcattactg gatctatcaa caggagtcca agcgagctcg taaacttggt ctgacagctc
     2041 tagctccggc aaaaaaacgg gcaaggtgtc accaccctgc cctttttctt taaaaccgaa
     2101 aagattactt cgcgtttgcc acctgacgtc taagaaaagg aatattcagc aatttgcccg
     2161 tgccgaagaa aggcccaccc gtgaaggtga gccagtgagt tgattgctac gtaattagtt
     2221 agttagccct tagtgactcg aattcaaaaa aatgcttgac aattaatcat cggctcgtat
     2281 aatgtgtgga attgtgagcg ctcacaattc cacacatcta gaaaggaagc taaaatggag
     2341 aaaaaaatca ctggatatac caccgttgat atatcccaat ggcatcgtaa agaacatttt
     2401 gaggcatttc agtcagttgc tcaatgtacc tataaccaga ccgttcagct ggatattacg
     2461 gcctttttaa agaccgtaaa gaaaaataag cacaagtttt atccggcctt tattcacatt
     2521 cttgcccgcc tgatgaatgc tcatccggag ttccgtatgg caatgaaaga cggtgagctg
     2581 gtgatatggg atagtgttca cccttgttac accgttttcc atgagcaaac tgaaacgttt
     2641 tcatcgctct ggagtgaata ccacgacgat ttccggcagt ttctacacat atattcgcaa
     2701 gatgtggcgt gttacggtga aaacctggcc tatttcccta aagggtttat tgagaatatg
     2761 tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt ttgatttaaa cgtggccaat
     2821 atggacaact tcttcgcccc cgttttcacc atgggcaaat attatacgca aggcgacaag
     2881 gtgctgatgc cgctggcgat tcaggttcat catgccgtct gtgatggctt ccatgtcggc
     2941 agaatgctta atgaattaca acagtactgc gatgagtggc agggcggggc gtaaaagctt
     3001 ttatttacct attaccttgg tgatctcgcc tttcacgtag tggacaaatt cttccaactg
     3061 atctgcgcgc gaggccaagc gatcttcttc ttgtccaaga taagcctgtc tagcttcaag
     3121 tatgacgggc tgatactggg ccggcaggcg ctccattgcc cagtcggcag cgacatcctt
     3181 cggcgcgatt ttgccggtta ctgcgctgta ccaaatgcgg gacaacgtaa gcactacatt
     3241 tcgctcatcg ccagcccagt cgggcggcga gttccatagc gttaaggttt catttagcgc
     3301 ctcaaataga tcctgttcag gaaccggatc aaagagttcc tccgccgctg gacctaccaa
     3361 ggcaacgcta tgttctcttg cttttgtcag caagatagcc agatcaatgt cgatcgtggc
     3421 tggctcgaag atacctgcaa gaatgtcatt gcgctgccat tctccaaatt gcagttcgcg
     3481 cttagctgga taacgccacg gaatgatgtc gtcgtgcaca acaatggtga cttctacagc
     3541 gcggagaatc tcgctctctc caggggaagc cgaagtttcc aaaaggtcgt tgatcaaagc
     3601 tcgccgcgtt gtttcatcaa gccttacggt caccgtaacc agcaaatcaa tatcactgtg
     3661 tggcttcagg ccgccatcca ctgcggagcc gtacaaatgt acggccagca acgtcggttc
     3721 gagatggcgc tcgatgacgc caactacctc tgatagttga gtcgatactt cggcgatcac
     3781 cgcttccctc atgatgttta actttgtttt agggcgactg ccctgctgcg taacatcgtt
     3841 gctgctccat aacatcaaac atcgacccac ggcgtaacgc gcttgctgct tggatgcccg
     3901 aggcatagac tgtaccccaa aaaaacagtc ataacaagcc atgaaaaccg ccactgcgcc
     3961 gttaccaccg ctgcgttcgg tcaaggttct ggaccagttg cgtaagcgca ttttagcttc
     4021 cttagctcct gaaaatctcg ataactcaaa aaatacgccc ggtagtgatc ttatttcatt
     4081 atggtgaaag ttggaacctc ttacctgcag gagtcactaa gggttagtta gttagattag
     4141 cagaaagtca aaagcctccg accggaggct tttgactaaa acttcccttg gggttatcat
     4201 tggggctcac tcaaaggcgg taatcagata aaaaaaatcc ttagctttcg ctaaggatga
     4261 tttctgctag t
//
