LOCUS       sequence-331373-        8584 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli Nissle pMUT1-derived plasmid with ampicillin resistance and 
            constitutive RFP expression and strong temperature sensitive GFP 
            expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8584)
  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 8584)
  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: pM1s3AsR_TS-C7
FEATURES             Location/Qualifiers
     source          1..8584
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(170..199)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     misc_feature    246..267
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     CDS             join(347..415,416..1207)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     terminator      complement(1337..1366)
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     promoter        1448..1482
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     RBS             1491..1502
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             1509..2186
                     /codon_start=1
                     /product="monomeric derivative of DsRed (Campbell et al., 
                     2002)"
                     /label=mRFP1
                     /translation="MASSEDVIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEGTQTAK
                     LKVTKGGPLPFAWDILSPQFQYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFEDGGV
                     VTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASTERMYPEDGALKGEIKM
                     RLKLKDGGHYDAEVKTTYMAKKPVQLPGAYKTDIKLDITSHNEDYTIVEQYERAEGRHS
                     TGA"
     terminator      2231..2302
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      2318..2345
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     misc_feature    2352..2372
                     /label=BioBrick suffix
                     /note="universal suffix for all parts"
     terminator      2408..2439
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     misc_RNA        2784..2833
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             2888..2905
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2918..3631
                     /codon_start=1
                     /product="green variant of teal fluorescent protein mTFP1 
                     (Ai et al., 2008)"
                     /label=mWasabi
                     /translation="MVSKGEETTMGVIKPDMKIKLKMEGNVNGHAFVIEGEGEGKPYDG
                     TNTINLEVKEGAPLPFSYDILTTAFSYGNRAFTKYPDDIPNYFKQSFPEGYSWERTMTF
                     EDKGIVKVKSDISMEEDSFIYEIHLKGENFPPNGPVMQKETTGWDASTERMYVRDGVLK
                     GDVKMKLLLEGGGHHRVDFKTIYRAKKAVKLPDYHFVDHRIEILNHDKDYNKVTVYEIA
                     VARNSTDGMDELYKG"
     promoter        3685..3762
                     /gene="lacI"
                     /label=lacI promoter
     CDS             4918..4962
                     /codon_start=1
                     /product="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
                     /label=S-Tag
                     /translation="KETAAAKFERQHMDS"
     terminator      5014..5061
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             5233..5250
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      5317..5364
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     7265..7284
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7507..7524
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 ctaaatctag aatgcggtcg agctcgtaaa cttggtctga cagctctagc tccggcaaaa
       61 aaacgggcaa ggtgtcacca ccctgccctt tttctttaaa accgaaaaga ttacttcgcg
      121 tttgccacct gacgtctaag aaaaggaata ttcagcaatt tgcccgtgcc gaagaaaggc
      181 ccacccgtga aggtgagcca gtgagttgat tgctacgtaa ttagttagtt agcccttagt
      241 gactcgaatt cgcggccgct tctagagcta aatacattca aatatctatc cgctcatgag
      301 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagaatatga gtattcaaca
      361 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
      421 agaaacgctg gtgaaagtaa aagatgccga agatcagttg ggtgcacgtg tgggttacat
      481 cgaactggac ctcaacagcg gtaagattct tgagagtttt cgccccgaag aacgtttccc
      541 aatgatgagc acttttaaag ttctgctctg tggcgcggta ttatcccgta ttgacgccgg
      601 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
      661 agtcacagaa aagcatctta cggacggcat gacagtacgc gaattatgca gcgctgccat
      721 aaccatgagt gataacacgg cggccaactt acttctgaca acgatcggag gaccgaagga
      781 gcttaccgct tttttgcaca acatgggtga tcatgtaact cgccttgatc gttgggaacc
      841 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagctatggc
      901 aacaacgttg cgcaaactct taactggcga acttcttact ctcgcttccc ggcaacaatt
      961 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     1021 tggctggttt attgctgata aatctggagc cggtgagcgt gggtcccgcg gtattattgc
     1081 agccctgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagcca
     1141 ggcaactatg gacgaacgta atcgccagat cgctgagata ggtgcctccc tgattaagca
     1201 ttggtaataa tactagctcc ggcaaaaaaa cgggcaaggt gtcaccaccc tgcccttttt
     1261 ctttaaaacc gaaaagatta cttcgcgttt gccacctgac gtctaagaaa aggaatattc
     1321 agcaatttgc ccgtgccgaa gaaaggccca cccgtgaagg tgagccagtg agttgattgc
     1381 tacgtaacaa tacgcaaacc gcctctcgtt ccttatcatc tggcgaatcg gacccacaag
     1441 agcactgttt acagctagct cagtcctagg tattatgcta gctactagag aaagaggaga
     1501 aatactagat ggcttcctcc gaagacgtta tcaaagagtt catgcgtttc aaagttcgta
     1561 tggaaggttc cgttaacggt cacgagttcg aaatcgaagg tgaaggtgaa ggtcgtccgt
     1621 acgaaggtac ccagaccgct aaactgaaag ttaccaaagg tggtccgctg ccgttcgctt
     1681 gggacatcct gtccccgcag ttccagtacg gttccaaagc ttacgttaaa cacccggctg
     1741 acatcccgga ctacctgaaa ctgtccttcc cggaaggttt caaatgggaa cgtgttatga
     1801 acttcgaaga cggtggtgtt gttaccgtta cccaggactc ctccctgcaa gacggtgagt
     1861 tcatctacaa agttaaactg cgtggtacca acttcccgtc cgacggtccg gttatgcaga
     1921 aaaaaaccat gggttgggaa gcttccaccg aacgtatgta cccggaagac ggtgctctga
     1981 aaggtgaaat caaaatgcgt ctgaaactga aagacggtgg tcactacgac gctgaagtta
     2041 aaaccaccta catggctaaa aaaccggttc agctgccggg tgcttacaaa accgacatca
     2101 aactggacat cacctcccac aacgaagact acaccatcgt tgaacagtac gaacgtgctg
     2161 aaggtcgtca ctccaccggt gcttaataac gctgatagtg ctagtgtaga tcgctactag
     2221 agccaggcat caaataaaac gaaaggctca gtcgaaagac tgggcctttc gttttatctg
     2281 ttgtttgtcg gtgaacgctc tctactagag tcacactggc tcaccttcgg gtgggccttt
     2341 ctgcgtttat atactagtag cggccgctgc aggagtcact aagggttagt tagttagatt
     2401 agcagaaagt caaaagcctc cgaccggagg cttttgacta aaacttccct tggggttatc
     2461 attggggctc actcaaaggc ggtaatcaga tgcggcgagc ggtatcagct cactcaaagg
     2521 cggtaatcag ataaaaaaaa tccttagctt tcgctaagga tgatttctgc tagtacgtac
     2581 ggtttcgaag cttgctcttc ttaagcatta ctcgcatcca ttctcaggct gtctcgtctc
     2641 gtctcctgac ctcctgccag caatagtaag acaacacgca aagtccgatc tcgatcccgc
     2701 gaaattttaa tttgtttgtt agttagttta tttgttggtt tgtttgtgtt atacaataag
     2761 cctctagaaa taattttgtt tagctgtcac cggatgtgct ttccggtctg atgagtccgt
     2821 gaggacgaaa cagcctctac aaattttgtt taaaacttta agaaggagat ataccatggg
     2881 cagcagccat caccatcatc accacagcca ggatccaatg gtgagcaagg gcgaggagac
     2941 cacaatgggc gtaatcaagc ccgacatgaa gatcaagctg aagatggagg gcaacgtgaa
     3001 tggccacgcc ttcgtgatcg agggcgaggg cgagggcaag ccctacgacg gcaccaacac
     3061 catcaacctg gaggtgaagg agggagcccc cctgcccttc tcctacgaca ttctgaccac
     3121 cgcgttcagt tacggcaaca gggccttcac caagtacccc gacgacatcc ccaactactt
     3181 caagcagtcc ttccccgagg gctactcttg ggagcgcacc atgaccttcg aggacaaggg
     3241 catcgtgaag gtgaagtccg acatctccat ggaggaggac tccttcatct acgagataca
     3301 cctcaagggc gagaacttcc cccccaacgg ccccgtgatg cagaaggaga ccaccggctg
     3361 ggacgcctcc accgagagga tgtacgtgcg cgacggcgtg ctgaagggcg acgtcaagat
     3421 gaagctgctg ctggagggcg gcggccacca ccgcgttgac ttcaagacca tctacagggc
     3481 caagaaggcg gtgaagctgc ccgactatca ctttgtggac caccgcatcg agatcctgaa
     3541 ccacgacaag gactacaaca aggtgaccgt ttacgagatc gccgtggccc gcaactccac
     3601 cgacggcatg gacgagctgt acaagggctg aaagcttgcg gccgcataat gcttaagtcg
     3661 aacagaaagt aatcgtatga gctcgacacc atcgaatggc gcaaaacctt tcgcggtatg
     3721 gcatgatagc gcccggaaga gagtcaattc agggtggtga atgtataaga aggagatata
     3781 catatgcgtc cggcgacata cgaaccagaa cagattattg aagcagggct ggccctgcag
     3841 gctgaaggac ggaatatcac cgggttcgca ctacgtaacc aggtgggtgg cggcaatccg
     3901 acacgtctcc gccagatatg ggacgaatac caggcttcac agagcacggt cgtcactgaa
     3961 ctcgttgccg agctgccagt ggaagtggct gaagaagtga aggccgtctc cgccgcgctg
     4021 tccgaacgca tcacccagct ggcgacagaa ctgaatgaca aggcggtccg ggctgcagaa
     4081 cgccgggttg cggaagtcac gcgtgctgcc ggtgaacaga ccgcacaggc agagcgggag
     4141 ctggccgacg ccgcgcagac agtcgacgac ctggaagaaa aactggttga actgcaggac
     4201 agatatgaca gtttgacgct ggcgctggag tcagaacgtt cactgcgtca gcagcatgat
     4261 gtggagatgg cccagctgaa agagcgtctt gcggccgctg aagagaatac ccgtcagcga
     4321 gaggaacggt atcaggagca gaggacagtg ctgcaggatg cgcttaatgc ggagcaggca
     4381 cagcacataa acacgcggga agaccagcag aaacgactgg agcaaatttc tgccgaagct
     4441 aatgcgcgta cagaagaact gaagtctgaa cgcgataaag tcaatactct ccttacccgc
     4501 cttgaatcgc aggaaaatgc gctggcctca gaacgtcagc agcatctggc cacccgcgaa
     4561 acgctgcagc aacgcctcga gcaggccatc gctgacacgc aggcgcgcgc cggtgagatt
     4621 gcacttgaac gtgacagagt cagcagcctc accgcaaggc tggaatcgca ggaaaaggcc
     4681 tcctcggagc aactggtgcg tatgggcagt gaaatagcca gtctgacaga gcgttgcaca
     4741 cagctggaaa accagcgtga tgatgcccgt ctggagacga tgggggagaa agaaacggtc
     4801 gcggcactgc gtggtgaggc tgaagccctg aagcgtcaga accagtcact gatggcggcg
     4861 ctttcaggca ataaacagac cggtggccag aatgcgtgag gtaccctcga gtctggtaaa
     4921 gaaaccgctg ctgcgaaatt tgaacgccag cacatggact cgtctactag cgcagcttaa
     4981 ttaacctagg ctgctgccac cgctgagcaa taactagcat aaccccttgg ggcctctaaa
     5041 cgggtcttga ggggtttttt gctgactcgg taccaaattt tcgaaaaaag acgctgaaaa
     5101 gcgtcttttt tcgttttggt ccgagccaac tccctttaca acctcactca agtccgttag
     5161 agccaggata catagattac cacaactccg agcccttcca cccgccggcg agacctctag
     5221 gccgcactcg agcaccacca ccaccaccac tgagatccgg ctgctaacaa agcccgaaag
     5281 gaagctgagt tggctgctgc caccgctgag caataactag cataacccct tggggcctct
     5341 aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggata aaccttgctt
     5401 cttcgcggtg tttcagtggt gcgtacaatt aagggattat ggtaaatata tgagctatgc
     5461 gataacttta actgtgaagc gatgaaccca ttacaggcaa agccaattac tcctgacagt
     5521 ggtttagcca gaagcagggc taccaagacc aatgcaataa gtaatatatc gttttgctat
     5581 cgtgcatcgc tcgcgctcag ttccattgtg cttttttaag ctgtcgtttt tcttacggta
     5641 tataccggtt ttttatggcg tggtttctta acttgttcag ctactgatgg acccattgta
     5701 tctaggtagt caactagctt tgttagatca taaatattgc gaccaccata tcggcgatca
     5761 ctcttcgatg ttggtttctt gtccaagaga ttagcttttt caagatcatt gatagctctc
     5821 tgaacagtcc gtacagaaac ccccatacgt atggctagac tttccattga cggatgcggc
     5881 cactcttgca aactccacca gtgaacgatc aggttaagta gtgtgttaaa ggccactgaa
     5941 gttagctttt tctcgttttg tataaaaaac aatacggtag gcactgctgt ccagccaaga
     6001 gacaaaccgc cagctttcca tttattctta acggagtaag tcattgattt tcctaagccc
     6061 caaaatattt aaagtatata ttatatggta tattcatatg aatagggtga cactggcgcc
     6121 attattgtgc aaccaaaaaa gactactctg aaaacgagga aaagattttt tcctgcctga
     6181 attagatacg gagttagcga tatgaaaacc gaacaacgtc atgatcttgt taaagatatt
     6241 gaggtttttg gcgtatcctt gtctctgttg atttccagag cgaatgagaa gtctgttaca
     6301 atgccatctg gtctaagtca ggagcagaga agagcatggg cagcggagca ggcgcgcaaa
     6361 atccacaatt gaatattgtc tcattctctg agaccttcaa cctttattac acatccagat
     6421 attctgcaaa aacactcgat aaaatcgatg atttcattga gcattttgaa aaatacaatc
     6481 tctttggcga tcctttaaaa ggatatccag cttggactgg caaagtatcg ccatcgtgga
     6541 aagtgcctga tcattacgaa aacaaagaag ctattgagaa gtatgctaga gctaacaaat
     6601 tatggcatgc tcatttaggc gatccggttt ttaaagatac gtttcatggg aaatacaagg
     6661 tttctgactg ggttattcat ttccagcggc tgacaccgaa ccatataaag cttttagagc
     6721 ttggatacca tgacccaatg aagctacctc aaaactttga atgatcgagc ggcaggctaa
     6781 atgaaatctt gagattcatt cagtctcgtc gtaatctcac tattgtaaaa acgaaaaaac
     6841 caccctggca ggtggttttt cgaaggttag ttaatcctgg cagattctct aaccgtggta
     6901 acagtcttgt gcgagacatg tcaccaaata ctgtcctttc agtgtagcct cagttaggcc
     6961 gccacttcaa gaactctcgt tacatctctc gcacatcctg cttaccagtg gccgttgcca
     7021 gtggcgttaa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     7081 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     7141 ccgaaccgag ataccaacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     7201 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     7261 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     7321 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc tcccgcgcgc
     7381 ccttcttctg ggtctttgtc ttttgctcac atgttctttc cggttttatc ccccgattct
     7441 gtggataacc gtattaccgc ctttgagtga gctgacaccg ctcgccgcag tcgaacgacc
     7501 gagcgtagcg agtcagtgag cgaggaagcg gaagagcgcc ttatgtgaca ttttctcctt
     7561 acgctcctct atgccgttct gcatcctgtc cggatgcgtt atatcccggt aagattttcc
     7621 gcttcaaagc gtgtctgtat gctgttctgg agttcttctg cgagttcgtg cagtttctca
     7681 cacatggcgg cctgttcgtc ggcattgagt gcgtccagtt tttcgagcag cgtcaggctc
     7741 tgacttttta tgaatcccgc catgttgagt acggcttgct gctgcttatt catcttttcg
     7801 ttttctccgt tctgtctgtc atctgcgttg tgtgattata tcgcgtacca cttttcgact
     7861 gttttgctgc cgctattctg ccgcttggct ttttgacggg catttctgtc agacaacact
     7921 gtcactgcca aaaaactgcc gtgcctttgt cggtaattcg agcttgctga caggacagga
     7981 tgtgcaattg ttataccgcg catacatgca cgctattaca attgccctgg tcagggcttt
     8041 gccccgacac ccatgtcaga tacggagcca tgttttatga caaaacgaag tggaagtaat
     8101 acgcgcaggc gggctatcag tcgccctgtt cgtctgacgg cagaagaaga ccaggaaatc
     8161 agaaaaaggg ctgctgaatg cggcaagacc gtttccggtt ttttacgggc ggcagctctc
     8221 ggtaagaaag ttaactcact gactgatgat cgggtactga aagaagttat gagactgggg
     8281 gcgttacaga aaaaactctt tatcgacggc aagcgtgtcg gggacaggga gtatgcggag
     8341 gtgctgatcg ctattacgga gtatcaccgt gccctgttat ccaggcttat ggcagattag
     8401 cttcccggag agaaaactgt cgaaaactga cggtatgaac accgtaagct cccaaagtga
     8461 tcaccattcg ctttcatgca tagctatgca gcgagctgaa aacgatcctg acgcatcctt
     8521 cctgtttttc cggggtaaaa catctctttt tgcggtgtct gcgtcagaat cgcgttcagc
     8581 gccc
//
