LOCUS       sequence-296678-        4221 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  T7 promoter driven synthesis of C-terminal fusion of sfGFP to TetR 
            repressor (TetR-sfGFP).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4221)
  AUTHORS   Niederholtmeyer H, Chaggan C, Devaraj NK
  TITLE     Communication and quorum sensing in non-living mimics of eukaryotic 
            cells.
  JOURNAL   Nat Commun. 2018 Nov 28;9(1):5027. doi: 10.1038/s41467-018-07473-7.
  PUBMED    30487584
REFERENCE   2  (bases 1 to 4221)
  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: "Nat 
            Commun."; date: "2018-11-28"; volume: "9(1):5027. doi"; pages: " 
            10.1038/s41467-018-07473-7"
COMMENT     Sequence Label: pTNT-pT7-tetR-sfGFP
FEATURES             Location/Qualifiers
     source          1..4221
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        35..53
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             110..121
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             128..748
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     complement(201..220)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     CDS             749..1462
                     /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"
     terminator      1530..1577
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1656..2111
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1743..1762)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1953..1974
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2178..2197)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2297..2319
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2357..2375)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2443..2547
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2548..2616,2617..3408)
                     /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"
     primer_bind     complement(2766..2785)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3579..4167
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4068..4087
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cacatggctc gacagatctt aaggctagag tacttaatac gactcactat aggctagcat
       61 ttaggtgaca ctataagcac atcagcagga cgcactgacc gaattcatta aagaggagaa
      121 aggtaccatg tccagattag ataaaagtaa agtgattaac agcgcattag agctgcttaa
      181 tgaggtcgga atcgaaggtt taacaacccg taaactcgcc cagaagctag gtgtagagca
      241 gcctacattg tattggcatg taaaaaataa gcgggctttg ctcgacgcct tagccattga
      301 gatgttagat aggcaccata ctcacttttg ccctttagaa ggggaaagct ggcaagattt
      361 tttacgtaat aacgctaaaa gttttagatg tgctttacta agtcatcgcg atggagcaaa
      421 agtacattta ggtacacggc ctacagaaaa acagtatgaa actctcgaaa atcaattagc
      481 ctttttatgc caacaaggtt tttcactaga gaatgcatta tatgcactca gcgctgtggg
      541 gcattttact ttaggttgcg tattggaaga tcaagagcat caagtcgcta aagaagaaag
      601 ggaaacacct actactgata gtatgccgcc attattacga caagctatcg aattatttga
      661 tcaccaaggt gcagagccag ccttcttatt cggccttgaa ttgatcatat gcggattaga
      721 aaaacaactt aaatgtgaaa gtgggtctat gcgtaaagga gaagaacttt tcactggtgt
      781 cgtccctatt ctggtggaac tggatggtga tgtcaacggt cataagtttt ccgtgcgtgg
      841 cgagggtgaa ggtgacgcaa ctaatggtaa actgacgctg aagttcatct gtactactgg
      901 taaactgccg gtaccttggc cgactctggt aacgacgctg acttatggtg ttcagtgctt
      961 tgctcgttat ccggaccata tgaagcagca tgacttcttc aagtccgcca tgccggaagg
     1021 ctatgtgcag gaacgcacga tttcctttaa ggatgacggc acgtacaaaa cgcgtgcgga
     1081 agtgaaattt gaaggcgata ccctggtaaa ccgcattgag ctgaaaggca ttgactttaa
     1141 agaagatggc aatatcctgg gccataagct ggaatacaat tttaacagcc acaatgttta
     1201 catcaccgcc gataaacaaa aaaatggcat taaagcgaat tttaaaattc gccacaacgt
     1261 ggaggatggc agcgtgcagc tggctgatca ctaccagcaa aacactccaa tcggtgatgg
     1321 tcctgttctg ctgccagaca atcactatct gagcacgcaa agcgttctgt ctaaagatcc
     1381 gaacgagaaa cgcgatcata tggttctgct ggagttcgta accgcagcgg gcatcacgca
     1441 tggtatggat gaactgtaca aataataatg gtacctctag agtcgacccg ggcggccgca
     1501 aaaaaaaaaa aaaaaaaaaa aaaaaaaaac tagcataacc ccttggggcc tctaaacggg
     1561 tcttgagggg ttttttggat ccgggctggc gtaatagcga agaggcccgc accgatcgcc
     1621 cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag
     1681 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     1741 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     1801 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     1861 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     1921 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     1981 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     2041 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     2101 gcttacaatt tcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     2161 atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     2221 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     2281 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     2341 cgcgcgagac gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata
     2401 atggtttctt agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     2461 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     2521 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     2581 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     2641 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     2701 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     2761 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     2821 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     2881 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     2941 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     3001 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     3061 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     3121 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     3181 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     3241 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     3301 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     3361 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     3421 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     3481 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     3541 tgagcgtcag accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc
     3601 gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat
     3661 caagagctac caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat
     3721 actgttcttc tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct
     3781 acatacctcg ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt
     3841 cttaccgggt tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg
     3901 gggggttcgt gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta
     3961 cagcgtgagc tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg
     4021 gtaagcggca gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg
     4081 tatctttata gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc
     4141 tcgtcagggg ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg
     4201 gccttttgct ggccttttgc t
//
