LOCUS       sequence-322897-        8783 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli Nissle pMUT2-derived plasmid with tetracycline resistance 
            and constitutive RFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8783)
  AUTHORS   Kan A, Gelfat I, Emani S, Praveschotinunt P, Joshi NS
  TITLE     Plasmid Vectors for in Vivo Selection-Free Use with the Probiotic E.
            coli Nissle 1917.
  JOURNAL   ACS Synth Biol. 2020 Dec 10. doi: 10.1021/acssynbio.0c00466.
  PUBMED    33301298
REFERENCE   2  (bases 1 to 8783)
  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: "ACS Synth 
            Biol. 2020 Dec 10. doi: 10.1021/acssynbio.0c00466."
COMMENT     Sequence Label: pM2s2TsR
FEATURES             Location/Qualifiers
     source          1..8783
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(4225..4254)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     misc_feature    4301..4322
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        join(4336..4341,4342..4358,4359..4364)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     CDS             4412..5602
                     /codon_start=1
                     /gene="tet"
                     /product="tetracycline efflux protein"
                     /label=TcR
                     /note="confers resistance to tetracycline"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVVGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     primer_bind     complement(4764..4783)
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     terminator      complement(5732..5761)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     promoter        5843..5877
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     RBS             5886..5897
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             5904..6581
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      6626..6697
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      6713..6740
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    6747..6767
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      6803..6834
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             7146..7163
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(7216..7234)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      7230..7277
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
ORIGIN
        1 caaaaaaccg acgataatca aaacaaacaa aaaattaact atatttgatg gtttgcttaa
       61 atcagtaaag accaacggca ttatgtacgt tgataaaaaa gaaagatact caccggattc
      121 ttttttacat gaaacgacct ttaactttct tgacaccgca ccggagtcta agtttttcaa
      181 aacccatcga taccaaatgt atgtataaga acaagttaaa atcaaagccc cgcagatcac
      241 tgacctcaat acagaaaatg ttaatctgct atttgaatag tcgagtacgc attgaaattt
      301 tccatccgcg ccagaacacg aagacatggc cttatctaaa acagaccata cgttatcaat
      361 accagaaaaa tatattgtta tcggtataaa taaaaacaac attgataaga gatacattct
      421 aattttcatt tttgtaaaat ttcctgtacc acgttgatct acttattcct aaagaaatcc
      481 attctccatc tctaactttc ggccttccac caccagagct tttttttcca cgttgacgct
      541 gaatttcaga agtatgtgtt tgtttaacat actcttcaaa gccaagctct gtaaggttct
      601 tacttgtcca cttagccaca cttttagcaa ttcccatgac ttcgttctcg tctaaaggtg
      661 cggagaactg caggttgtag gctttagcgc gttcaatgca ggcttgtagc cattggtcat
      721 actgcggcca gccttggcgg atagcgcggt aagcccactt gcgggtttta tcgaagaggg
      781 tgcagttacg gcctaaaccg tagtccggca ggatttcgcg gtcattggct gcgccaaggt
      841 cgaggtaatc ggctaaccag tcaagggtat agagctctgg ctgccagacg gtgatctgcc
      901 agtgcaggtg gttcggattc ttgcaaatta gccctgaata ccccgcatct gcgcccaatt
      961 ttttacgcag cgcattctcg atggcggcgg cgtatttaag gggggcagct cgaccatccg
     1021 gcgcggtacg taccgccgta tgcaaggcat acaacaggtg agcatgtccg ttctcggggt
     1081 ttttgatggt gagtgtgggc gcaggtgccc ccagatcggc ccaatcaatc gcggctccgg
     1141 ctctgtccac gtcaaagcaa agccagtaca tggcgtgagg ctgattaaac tggatgtatt
     1201 ttgcgaggag agcacgctct ttaccggcaa tgcgaacacc aaactgtaaa tcatcggaga
     1261 agtacggctt gtggggtaac cggtcgttaa aaagcgttaa agcctgatta tccaaggctc
     1321 ccagccttat ggcggggctg ttgttttgca cgctgcatgt gctaatatcc tttctaggtt
     1381 tcgacctagc cctgaatgtc atgtccgctc gccaaagtag agcatgattt cggggctttg
     1441 ttttttctgc cactaagtta cacctcaaca acggtttttg tcatccccga caatccgtta
     1501 ttcctgcttg ttctcgcacg gctttacgct catactactt cttgtagata cacttgtcac
     1561 tacatcaaga ggtgagatga tggccacgat taatattcgg atcgatgacg agctgaaaag
     1621 ccgctcttat gccgcactgg aaaagctggg cgtaacgccg tccgaggttc tgcgccaaac
     1681 actggaatat gtggcccaaa gcggacgttt gccgttccag caggttttgc tgaccgagga
     1741 tgatgccgat ttgatggcta tcgttcggga tcgtctggaa aacccacagg cgggcgtaaa
     1801 ggtgtcactg gatgagctat aaccttgaat ttgatccccg agccctgaag gaatggcgca
     1861 agctcgggga tgatgtccgt ctgcagttca agaaaaaact cgagcaggtt ctacaacacc
     1921 cgcggatcga taaaaatcgc ctgcgagagc tgcatgactg ctacaaaatc aagctccgtg
     1981 catccggtta tcgcttggtc tatcaggttc gcgatcaaac cattacggta ttcgtggtgg
     2041 cggtcggtaa gcgcgagcgt tctgccgctt acgatgcggc cgataaacgc ttataaactc
     2101 atgccgtgac cgcgagaata ccgctgttcg tgccgcttgg ctaattgctc aagcggcgca
     2161 ctgttctgtt taagctctcg acttgctgcg ccaagccggt gacttgcatg ctcaaggcct
     2221 gacaaaccct gtcgtttttc gccaactcct gcgaggtaac ctcgaacatg cgctgtaagt
     2281 tggcgtagct gtcctgccac gcttgctgct gttgttcgta gtgcctctgt aagctctcta
     2341 atgcgctcag aagctgctgc tccatttcgg tcatgaatct cttcaccctg atagataaaa
     2401 ccgcccagaa tcgattctgt ggcgtctgat gaggttattt ggcgctgtac ttgatgacct
     2461 gacgatgttg agcgttcttg tactcgtcga tcttcttcgc cccctgcgga aggatcaggt
     2521 aatacacgct cttgttcttg gaatcgtgaa ttatcgatac gccggctccg gtctggctct
     2581 ttaagtcctg caggatctgg ctctgctcgc tgatttcgtt ctggcgttcg accacgatag
     2641 tcccgagata ccaagctact ccaatcaata tcgcaaacag gatcccactt aatgccaggc
     2701 tgtacagcca tgtcattccg actaagcggt gtatctgttt tagctggctg tcgttctctt
     2761 cttggatagc ggtctgtatg ttccctgagc ttaatttcaa ggcctcggtg atacgtttct
     2821 cgtgcttctc gaatgcgttc gcgacgctcg ttgcggtagt cttggcttgc tgcttcgatt
     2881 tgctctcgaa ctcccgcgct aaatttaaaa tctcgctcat acagcactcc ttttaagcga
     2941 atattcgggc cacctgccgg atcagcaata ctgatactgg atttggtttc ccgtacgacc
     3001 gacaatccgg catcggtaag gtgggaaatc acccctttac gatccgtaat ttctccctgc
     3061 tcaatcaagc tgattagccc tttggtaatg gcttccgctg cctgctgttt gttgcgagga
     3121 aggtcattag agggggttaa tgctcggcga ttagcagggt cattcgggtc gcgtaaccca
     3181 agccggtcat tggtgagggt ttgccatgcg ttaacacgag gccggtcagc ccgatcaaag
     3241 taaggttgta gccgttttcc gctctgcaat tcgatgttcg ggataacaaa attcaattca
     3301 agacgccctt tgtcctgatg ttgaacccag aggcaggcat actggtcttt atctagaccg
     3361 gtcatcaatg tctgctccca ttcatccatc aatcgctgct tttcgccttc gggtaaatca
     3421 ctctcctgaa aagagagcac gccagaggta taagttcggg caaattcgca gccatcaatc
     3481 agctctttga cgtgctcagg gttaccccgt aacaccgtcg cttgttcgcg ctgacgatca
     3541 gggcccagaa ggtaatcgac aggaccactc ccgccaccgg caccacgacc atgaatctta
     3601 acgatcacga tgttgctcca gcagttcggc aagatgttgg tcaatgctat tgagcaccgc
     3661 taacaacgac acccgttctt gcggcggtaa gccatgctga ttcaagtaac gggctatttg
     3721 attgaggtta ttaccgatcc cgctgacctg acgtaacaag gtcgggtcta cggtaggtag
     3781 cggacgacgc cgagctgtac gcgattcgcc taagccaacg gctcgtaacc actcggccaa
     3841 atgcttacgg tcacagcgtt caagtagccg ctgatgctcc gcttcggtga gtctgatttt
     3901 gatctctttg gtgcgctttt ccatgagaat ccgctgagaa agtttcgcac ccaaagtgcg
     3961 aattttcgca gtggatgcaa ggggtttcgg ggggcggcga gccccctgaa acagtcacag
     4021 acggcacctc gaagagggga cgctgtgtgt actccctaaa tctagaatgc ggtcgagctc
     4081 gtaaacttgg tctgacagct ctagctccgg caaaaaaacg ggcaaggtgt caccaccctg
     4141 ccctttttct ttaaaaccga aaagattact tcgcgtttgc cacctgacgt ctaagaaaag
     4201 gaatattcag caatttgccc gtgccgaaga aaggcccacc cgtgaaggtg agccagtgag
     4261 ttgattgcta cgtaattagt tagttagccc ttagtgactc gaattcgcgg ccgcttctag
     4321 aggagattct catgtttgac agcttatcat cgataagctt taatgcggta gtttatcaca
     4381 gttaaattgc taacgcagtc aggcaccgtg tatgaaatct aacaatgcgc tcatcgtcat
     4441 tctcggcacc gtcaccctgg acgctgtagg cataggcttg gttatgccgg tactgccggg
     4501 cctcttgcgg gatatcgtcc attccgacag tattgccagt cactatggcg tgctgcttgc
     4561 gctctatgcg ttgatgcaat ttctttgcgc acccgttctc ggagccctgt ccgaccgctt
     4621 tggccgccgt ccagtcctgc tcgcttcgct ccttggagcc actatcgact acgcgatcat
     4681 ggcgaccaca cccgtcctgt ggattctcta cgccggacgc atcgtggcgg gcatcacggg
     4741 tgccacaggt gcggttgctg gtgcctatat cgccgacatc accgatgggg aagatcgggc
     4801 tcgccacttc gggctcatga gcgcttgttt cggcgtgggt atggtggtag gccccgtggc
     4861 cgggggactg ttgggtgcca tctccttgca tgcaccattc cttgcggcgg cggtgctcaa
     4921 cggcctcaac ctcctcctgg gctgcttcct tatgcaggaa tcgcataagg gagagcgccg
     4981 tccgatgccc ttgcgtgcct tcaatccagt cagctccttc cggtgggcgc ggggcatgac
     5041 tatcgtcgcc gcacttatga ctgttttctt tatcatgcaa ctcgtaggac aggttccggc
     5101 agcgctctgg gtcattttcg gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct
     5161 gtcgcttgcg gtattcggaa tcttgcacgc cctcgctcaa gccttcgtca cgggccccgc
     5221 caccaaacgt ttcggcgaga agcaggccat tatcgcgggc atggcggccg acgcgctggg
     5281 ctacgtcttg ctggcgttcg cgacgcgcgg ctggatggcc ttccccatta tgattcttct
     5341 cgcttccggc ggcatcggta tgcccgcgtt gcaggccatg ctgtcccgcc aagtagatga
     5401 cgaccatcag ggacagcttc aagggtcgct cgcggctctt accagcctca cttcgatcat
     5461 tggaccgctg atcgtcacgg cgatttatgc cgcctcggcg agcacatgga acgggttggc
     5521 atggattgta ggtgccgccc tttaccttgt ctgcctcccc gcgttgcgtc gcggtgcatg
     5581 gagccgggcc acctcgacct aataatacta gctccggcaa aaaaacgggc aaggtgtcac
     5641 caccctgccc tttttcttta aaaccgaaaa gattacttcg cgtttgccac ctgacgtcta
     5701 agaaaaggaa tattcagcaa tttgcccgtg ccgaagaaag gcccacccgt gaaggtgagc
     5761 cagtgagttg attgctacgt aacaatacgc aaaccgcctc tcgttcctta tcatctggcg
     5821 aatcggaccc acaagagcac tgtttacagc tagctcagtc ctaggtatta tgctagctac
     5881 tagagaaaga ggagaaatac tagatggctt cctccgaaga cgttatcaaa gagttcatgc
     5941 gtttcaaagt tcgtatggaa ggttccgtta acggtcacga gttcgaaatc gaaggtgaag
     6001 gtgaaggtcg tccgtacgaa ggtacccaga ccgctaaact gaaagttacc aaaggtggtc
     6061 cgctgccgtt cgcttgggac atcctgtccc cgcagttcca gtacggttcc aaagcttacg
     6121 ttaaacaccc ggctgacatc ccggactacc tgaaactgtc cttcccggaa ggtttcaaat
     6181 gggaacgtgt tatgaacttc gaagacggtg gtgttgttac cgttacccag gactcctccc
     6241 tgcaagacgg tgagttcatc tacaaagtta aactgcgtgg taccaacttc ccgtccgacg
     6301 gtccggttat gcagaaaaaa accatgggtt gggaagcttc caccgaacgt atgtacccgg
     6361 aagacggtgc tctgaaaggt gaaatcaaaa tgcgtctgaa actgaaagac ggtggtcact
     6421 acgacgctga agttaaaacc acctacatgg ctaaaaaacc ggttcagctg ccgggtgctt
     6481 acaaaaccga catcaaactg gacatcacct cccacaacga agactacacc atcgttgaac
     6541 agtacgaacg tgctgaaggt cgtcactcca ccggtgctta ataacgctga tagtgctagt
     6601 gtagatcgct actagagcca ggcatcaaat aaaacgaaag gctcagtcga aagactgggc
     6661 ctttcgtttt atctgttgtt tgtcggtgaa cgctctctac tagagtcaca ctggctcacc
     6721 ttcgggtggg cctttctgcg tttatatact agtagcggcc gctgcaggag tcactaaggg
     6781 ttagttagtt agattagcag aaagtcaaaa gcctccgacc ggaggctttt gactaaaact
     6841 tcccttgggg ttatcattgg ggctcactca aaggcggtaa tcagatgcgg cgagcggtat
     6901 cagctcactc aaaggcggta atcagataaa aaaaatcctt agctttcgct aaggatgatt
     6961 tctgctagta cgtacggttt cgaagcttgc tcttcttaag cattactcgc atccattctc
     7021 aggctgtctc gtctcgtctc cgcgcgcggt cttcctggag atgatcaaac ctaggccagg
     7081 atacatagat taccacaact ccgagccctt ccacccgccg gcgagacctc taggccgcac
     7141 tcgagcacca ccaccaccac cactgagatc cggctgctaa caaagcccga aaggaagctg
     7201 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     7261 tcttgagggg ttttttgctg aaaggaggaa ctatatccgg ataaaccttg cttcttcgcg
     7321 gtggcttagt acagcatcta tcgtacatcg aggtcgcatc acgcacaaac aaaaagcccc
     7381 gcaaaagcag ggctgttatc tgatataggt tgttttgtct cacacggcag cggaagagga
     7441 atccgaagtg gtactggtag tagtattgga tgctgctgac gatattttcc gctttgaccc
     7501 aaggcttaaa taatcaatgc ctgtaatcaa cgatctcaat acgccttcgg ataccatagc
     7561 gataaacgta tcttgctggt tatggcttgc gatgcaaatc gtagcatcac cttttttata
     7621 ctttaaaaca cctgctaaat atccattttc atctagaaca ctcttaagat gttcatttgt
     7681 tattgtttgt agcgtttgct ttgtttcgct tcgagcatac gccttagcta gcttccgaga
     7741 aaaagcatcc gcatcatgac tatctttatt tactcgctca ataaatttgc ttaagtcaac
     7801 aaatccctta aaacgagtgg acatattgtt aacaaaatca gtggcagcat tttttatcca
     7861 tgctttatag ccaaaaaaac gctcgaaaac attttggtcg tagataaata ccgtatcgcc
     7921 agcaaaaaca agagatgcct taccatcaat agaaatcata tcttgatcta ctcgagatag
     7981 ttcttttttg ctaaacccat aaaaattatt tttctttctt gataagtttt gaactggata
     8041 ttgcttcttg tatatggtta tacaattgtc gtcattattc ttactcaaaa cgaaaatgat
     8101 tgagtcaact tttgatatta gatccacact gtcaaaatca acaattggga tattttcatt
     8161 gccatcacca ccaaacccag catcaaacac aatgccagtc agaaacttca tttgctcatt
     8221 atcatactgc tcagcatcat ttaaaaatga aatggtgttg gtgcggtcac ttagtgtatt
     8281 aacatctgat actatcacaa cccgattttc taaatcagat atagttttct ttatcacatt
     8341 atcaataatg gactgtttta gctcactgtc atttttaagg atggcaattt tatagctaaa
     8401 agagtcctta gcacccgctt tacctttatt tttaaagtta aagtaagtgt gcaatgtaac
     8461 atcgttaata tcacaatcaa aatgcttata cagttctaaa agctcttgtg ctttttcttc
     8521 attatgctcc aaagcatcaa gatctaaggc atcgtcactc atcatcattc ctctatgatt
     8581 tttttcgcga acgttaaata atcatcatga tttataactc tgataaaatc attttctttt
     8641 attaaatctt tagataaaac tatcaactca ccgtcttgcg ttttttccct tccattagct
     8701 accacactgt aagtaatctt ataggcagaa acattaaata atgacaatgt tgggttgcag
     8761 tgaattcttt ttgttttgat gtg
//
