LOCUS       sequence-331367-        8358 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E. coli Nissle pMUT2-derived plasmid with ampicillin resistance and 
            constitutive GFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8358)
  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 8358)
  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: pM2s2AsG
FEATURES             Location/Qualifiers
     source          1..8358
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     RBS             463..474
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        complement(483..517)
                     /label=BBa_J23101
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.7"
     terminator      599..628
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     terminator      complement(688..746)
                     /label=his operon terminator
                     /note="This putative transcriptin terminator from the E. 
                     coli his operon has a 2-bp deletion introduced during 
                     synthesis. Its efficiency has not been determined."
     CDS             complement(join(757..1548,1549..1617))
                     /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"
     misc_feature    1697..1718
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     terminator      1765..1794
                     /label=T3Te terminator
                     /note="phage T3 early transcription terminator"
     terminator      complement(7525..7572)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     primer_bind     7568..7586
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     CDS             complement(7639..7656)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      complement(7939..7966)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(7982..8053)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(join(8101..8358,1..456))
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="MRKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFARYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
ORIGIN
        1 gatgtaaaca ttgtggctgt taaaattgta ttccagctta tggcccagga tattgccgtc
       61 ttctttaaag tcaatgcctt tcagctcaat gcggtttacc agggtatcgc cttcaaattt
      121 cacttccgca cgcgttttgt acgtgccgtc atccttaaag gaaatcgtgc gttcctgcac
      181 atagccttcc ggcatggcgg acttgaagaa gtcatgctgc ttcatatggt ccggataacg
      241 agcaaagcac tgaacaccat aagtcagcgt cgttaccaga gtcggccaag gtaccggcag
      301 tttaccagta gtacagatga acttcagcgt cagtttacca ttagttgcgt caccttcacc
      361 ctcgccacgc acggaaaact tatgaccgtt gacatcacca tccagttcca ccagaatagg
      421 gacgacacca gtgaacagct cttcgccttt acgcatctag tatttctcct ctttctctag
      481 tagctagcat aatacctagg actgagctag ctgtaaacag tgctcttgtg ggtccgattc
      541 gccagatgat aaggaacgag aggcggtttg cgtattgtta cgtagcaatc aactcactgg
      601 ctcaccttca cgggtgggcc tttcttcggc acgggcaaat tgctgaatat tccttttctt
      661 agacgtcagg tggcaaacgc gaagtaatct tttcggtttt aaagaaaaag ggcagggtgg
      721 tgacaccttg cccgtttttt gccggagcta gtattattac caatgcttaa tcagggaggc
      781 acctatctca gcgatctggc gattacgttc gtccatagtt gcctggctcc ccgtcgtgta
      841 gataactacg atacgggagg gcttaccatc tggccccagg gctgcaataa taccgcggga
      901 cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
      961 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     1021 gagagtaaga agttcgccag ttaagagttt gcgcaacgtt gttgccatag ctacaggcat
     1081 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     1141 gcgagttaca tgatcaccca tgttgtgcaa aaaagcggta agctccttcg gtcctccgat
     1201 cgttgtcaga agtaagttgg ccgccgtgtt atcactcatg gttatggcag cgctgcataa
     1261 ttcgcgtact gtcatgccgt ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     1321 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     1381 taataccgcg ccacagagca gaactttaaa agtgctcatc attgggaaac gttcttcggg
     1441 gcgaaaactc tcaagaatct taccgctgtt gaggtccagt tcgatgtaac ccacacgtgc
     1501 acccaactga tcttcggcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     1561 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatatt
     1621 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatagat
     1681 atttgaatgt atttagctct agaagcggcc gcgaattcga gtcactaagg gctaactaac
     1741 taattacgta gcaatcaact cactggctca ccttcacggg tgggcctttc ttcggcacgg
     1801 gcaaattgct gaatattcct tttcttagac gtcaggtggc aaacgcgaag taatcttttc
     1861 ggttttaaag aaaaagggca gggtggtgac accttgcccg tttttttgcc ggagctagag
     1921 ctgtcagacc aagtttacga gctcgaccgc attctagatt tagggagtac acacagcgtc
     1981 ccctcttcga ggtgccgtct gtgactgttt cagggggctc gccgcccccc gaaacccctt
     2041 gcatccactg cgaaaattcg cactttgggt gcgaaacttt ctcagcggat tctcatggaa
     2101 aagcgcacca aagagatcaa aatcagactc accgaagcgg agcatcagcg gctacttgaa
     2161 cgctgtgacc gtaagcattt ggccgagtgg ttacgagccg ttggcttagg cgaatcgcgt
     2221 acagctcggc gtcgtccgct acctaccgta gacccgacct tgttacgtca ggtcagcggg
     2281 atcggtaata acctcaatca aatagcccgt tacttgaatc agcatggctt accgccgcaa
     2341 gaacgggtgt cgttgttagc ggtgctcaat agcattgacc aacatcttgc cgaactgctg
     2401 gagcaacatc gtgatcgtta agattcatgg tcgtggtgcc ggtggcggga gtggtcctgt
     2461 cgattacctt ctgggccctg atcgtcagcg cgaacaagcg acggtgttac ggggtaaccc
     2521 tgagcacgtc aaagagctga ttgatggctg cgaatttgcc cgaacttata cctctggcgt
     2581 gctctctttt caggagagtg atttacccga aggcgaaaag cagcgattga tggatgaatg
     2641 ggagcagaca ttgatgaccg gtctagataa agaccagtat gcctgcctct gggttcaaca
     2701 tcaggacaaa gggcgtcttg aattgaattt tgttatcccg aacatcgaat tgcagagcgg
     2761 aaaacggcta caaccttact ttgatcgggc tgaccggcct cgtgttaacg catggcaaac
     2821 cctcaccaat gaccggcttg ggttacgcga cccgaatgac cctgctaatc gccgagcatt
     2881 aaccccctct aatgaccttc ctcgcaacaa acagcaggca gcggaagcca ttaccaaagg
     2941 gctaatcagc ttgattgagc agggagaaat tacggatcgt aaaggggtga tttcccacct
     3001 taccgatgcc ggattgtcgg tcgtacggga aaccaaatcc agtatcagta ttgctgatcc
     3061 ggcaggtggc ccgaatattc gcttaaaagg agtgctgtat gagcgagatt ttaaatttag
     3121 cgcgggagtt cgagagcaaa tcgaagcagc aagccaagac taccgcaacg agcgtcgcga
     3181 acgcattcga gaagcacgag aaacgtatca ccgaggcctt gaaattaagc tcagggaaca
     3241 tacagaccgc tatccaagaa gagaacgaca gccagctaaa acagatacac cgcttagtcg
     3301 gaatgacatg gctgtacagc ctggcattaa gtgggatcct gtttgcgata ttgattggag
     3361 tagcttggta tctcgggact atcgtggtcg aacgccagaa cgaaatcagc gagcagagcc
     3421 agatcctgca ggacttaaag agccagaccg gagccggcgt atcgataatt cacgattcca
     3481 agaacaagag cgtgtattac ctgatccttc cgcagggggc gaagaagatc gacgagtaca
     3541 agaacgctca acatcgtcag gtcatcaagt acagcgccaa ataacctcat cagacgccac
     3601 agaatcgatt ctgggcggtt ttatctatca gggtgaagag attcatgacc gaaatggagc
     3661 agcagcttct gagcgcatta gagagcttac agaggcacta cgaacaacag cagcaagcgt
     3721 ggcaggacag ctacgccaac ttacagcgca tgttcgaggt tacctcgcag gagttggcga
     3781 aaaacgacag ggtttgtcag gccttgagca tgcaagtcac cggcttggcg cagcaagtcg
     3841 agagcttaaa cagaacagtg cgccgcttga gcaattagcc aagcggcacg aacagcggta
     3901 ttctcgcggt cacggcatga gtttataagc gtttatcggc cgcatcgtaa gcggcagaac
     3961 gctcgcgctt accgaccgcc accacgaata ccgtaatggt ttgatcgcga acctgataga
     4021 ccaagcgata accggatgca cggagcttga ttttgtagca gtcatgcagc tctcgcaggc
     4081 gatttttatc gatccgcggg tgttgtagaa cctgctcgag ttttttcttg aactgcagac
     4141 ggacatcatc cccgagcttg cgccattcct tcagggctcg gggatcaaat tcaaggttat
     4201 agctcatcca gtgacacctt tacgcccgcc tgtgggtttt ccagacgatc ccgaacgata
     4261 gccatcaaat cggcatcatc ctcggtcagc aaaacctgct ggaacggcaa acgtccgctt
     4321 tgggccacat attccagtgt ttggcgcaga acctcggacg gcgttacgcc cagcttttcc
     4381 agtgcggcat aagagcggct tttcagctcg tcatcgatcc gaatattaat cgtggccatc
     4441 atctcacctc ttgatgtagt gacaagtgta tctacaagaa gtagtatgag cgtaaagccg
     4501 tgcgagaaca agcaggaata acggattgtc ggggatgaca aaaaccgttg ttgaggtgta
     4561 acttagtggc agaaaaaaca aagccccgaa atcatgctct actttggcga gcggacatga
     4621 cattcagggc taggtcgaaa cctagaaagg atattagcac atgcagcgtg caaaacaaca
     4681 gccccgccat aaggctggga gccttggata atcaggcttt aacgcttttt aacgaccggt
     4741 taccccacaa gccgtacttc tccgatgatt tacagtttgg tgttcgcatt gccggtaaag
     4801 agcgtgctct cctcgcaaaa tacatccagt ttaatcagcc tcacgccatg tactggcttt
     4861 gctttgacgt ggacagagcc ggagccgcga ttgattgggc cgatctgggg gcacctgcgc
     4921 ccacactcac catcaaaaac cccgagaacg gacatgctca cctgttgtat gccttgcata
     4981 cggcggtacg taccgcgccg gatggtcgag ctgcccccct taaatacgcc gccgccatcg
     5041 agaatgcgct gcgtaaaaaa ttgggcgcag atgtggggta ttcagggcta atttgcaaga
     5101 atccgaacca cctgcactgg cagatcaccg tctggcagcc agagctctat acccttgact
     5161 ggttagccga ttacctcgac cttggcgcag ccaatgaccg cgaaatcctg ccggactacg
     5221 gtttaggccg taactgcacc ctcttcgata aaacccgcaa gtgggcttac cgcgctatcc
     5281 gccaaggctg gccgcagtat gaccaatggc tacaagcctg cattgaacgc gctaaagcct
     5341 acaacctgca gttctccgca cctttagacg agaacgaagt catgggaatt gctaaaagtg
     5401 tggctaagtg gacaagtaag aaccttacag agcttggctt tgaagagtat gttaaacaaa
     5461 cacatacttc tgaaattcag cgtcaacgtg gaaaaaaaag ctctggtggt ggaaggccga
     5521 aagttagaga tggagaatgg atttctttag gaataagtag atcaacgtgg tacaggaaat
     5581 tttacaaaaa tgaaaattag aatgtatctc ttatcaatgt tgtttttatt tataccgata
     5641 acaatatatt tttctggtat tgataacgta tggtctgttt tagataaggc catgtcttcg
     5701 tgttctggcg cggatggaaa atttcaatgc gtactcgact attcaaatag cagattaaca
     5761 ttttctgtat tgaggtcagt gatctgcggg gctttgattt taacttgttc ttatacatac
     5821 atttggtatc gatgggtttt gaaaaactta gactccggtg cggtgtcaag aaagttaaag
     5881 gtcgtttcat gtaaaaaaga atccggtgag tatctttctt ttttatcaac gtacataatg
     5941 ccgttggtct ttactgattt aagcaaacca tcaaatatag ttaatttttt gtttgttttg
     6001 attatcgtcg gttttttgca catcaaaaca aaaagaattc actgcaaccc aacattgtca
     6061 ttatttaatg tttctgccta taagattact tacagtgtgg tagctaatgg aagggaaaaa
     6121 acgcaagacg gtgagttgat agttttatct aaagatttaa taaaagaaaa tgattttatc
     6181 agagttataa atcatgatga ttatttaacg ttcgcgaaaa aaatcataga ggaatgatga
     6241 tgagtgacga tgccttagat cttgatgctt tggagcataa tgaagaaaaa gcacaagagc
     6301 ttttagaact gtataagcat tttgattgtg atattaacga tgttacattg cacacttact
     6361 ttaactttaa aaataaaggt aaagcgggtg ctaaggactc ttttagctat aaaattgcca
     6421 tccttaaaaa tgacagtgag ctaaaacagt ccattattga taatgtgata aagaaaacta
     6481 tatctgattt agaaaatcgg gttgtgatag tatcagatgt taatacacta agtgaccgca
     6541 ccaacaccat ttcattttta aatgatgctg agcagtatga taatgagcaa atgaagtttc
     6601 tgactggcat tgtgtttgat gctgggtttg gtggtgatgg caatgaaaat atcccaattg
     6661 ttgattttga cagtgtggat ctaatatcaa aagttgactc aatcattttc gttttgagta
     6721 agaataatga cgacaattgt ataaccatat acaagaagca atatccagtt caaaacttat
     6781 caagaaagaa aaataatttt tatgggttta gcaaaaaaga actatctcga gtagatcaag
     6841 atatgatttc tattgatggt aaggcatctc ttgtttttgc tggcgatacg gtatttatct
     6901 acgaccaaaa tgttttcgag cgtttttttg gctataaagc atggataaaa aatgctgcca
     6961 ctgattttgt taacaatatg tccactcgtt ttaagggatt tgttgactta agcaaattta
     7021 ttgagcgagt aaataaagat agtcatgatg cggatgcttt ttctcggaag ctagctaagg
     7081 cgtatgctcg aagcgaaaca aagcaaacgc tacaaacaat aacaaatgaa catcttaaga
     7141 gtgttctaga tgaaaatgga tatttagcag gtgttttaaa gtataaaaaa ggtgatgcta
     7201 cgatttgcat cgcaagccat aaccagcaag atacgtttat cgctatggta tccgaaggcg
     7261 tattgagatc gttgattaca ggcattgatt atttaagcct tgggtcaaag cggaaaatat
     7321 cgtcagcagc atccaatact actaccagta ccacttcgga ttcctcttcc gctgccgtgt
     7381 gagacaaaac aacctatatc agataacagc cctgcttttg cggggctttt tgtttgtgcg
     7441 tgatgcgacc tcgatgtacg atagatgctg tactaagcca ccgcgaagaa gcaaggttta
     7501 tccggatata gttcctcctt tcagcaaaaa acccctcaag acccgtttag aggccccaag
     7561 gggttatgct agttattgct cagcggtggc agcagccaac tcagcttcct ttcgggcttt
     7621 gttagcagcc ggatctcagt ggtggtggtg gtggtgctcg agtgcggcct agaggtctcg
     7681 ccggcgggtg gaagggctcg gagttgtggt aatctatgta tcctggccta ggtttgatca
     7741 tctccaggaa gaccgcgcgc ggagacgaga cgagacagcc tgagaatgga tgcgagtaat
     7801 gcttaagaag agcaagcttc gaaaccgtac gtactagcag aaatcatcct tagcgaaagc
     7861 taaggatttt ttttatctga ttaccgcctt tgagtgagct gataccgctc gccgcagcgg
     7921 ccgctactag tatataaacg cagaaaggcc cacccgaagg tgagccagtg tgactctagt
     7981 agagagcgtt caccgacaaa caacagataa aacgaaaggc ccagtctttc gactgagcct
     8041 ttcgttttat ttgatgcctg gctctagtag cgatctacac tagcactatc agcgttatca
     8101 tttgtacagt tcatccatac catgcgtgat gcccgctgcg gttacgaact ccagcagaac
     8161 catatgatcg cgtttctcgt tcggatcttt agacagaacg ctttgcgtgc tcagatagtg
     8221 attgtctggc agcagaacag gaccatcacc gattggagtg ttttgctggt agtgatcagc
     8281 cagctgcacg ctgccatcct ccacgttgtg gcgaatttta aaattcgctt taatgccatt
     8341 tttttgttta tcggcggt
//
