LOCUS       sequence-296668-        4206 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  T7 promoter driven synthesis of TetR-mCherry fusion protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4206)
  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 4206)
  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-mCherry
FEATURES             Location/Qualifiers
     source          1..4206
                     /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..1459
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(894..912)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1162..1181
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     terminator      1515..1562
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1641..2096
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1728..1747)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1938..1959
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2163..2182)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2282..2304
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2342..2360)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2428..2532
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2533..2601,2602..3393)
                     /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(2751..2770)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3564..4152
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4053..4072
                     /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 ggtgagcaag ggcgaggagg ataacatggc
      781 catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga acggccacga
      841 gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct
      901 gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc ctcagttcat
      961 gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact tgaagctgtc
     1021 cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gcgtggtgac
     1081 cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga agctgcgcgg
     1141 caccaacttc ccctccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc
     1201 ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc agaggctgaa
     1261 gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg ccaagaagcc
     1321 cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct cccacaacga
     1381 ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca ccggcggcat
     1441 ggacgagctg tacaagtaat aatggtacct ctagagtcga cccgggcggc cgcaaaaaaa
     1501 aaaaaaaaaa aaaactagca taaccccttg gggcctctaa acgggtcttg aggggttttt
     1561 tggatccggg ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc
     1621 gcagcctgaa tggcgaatgg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
     1681 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
     1741 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
     1801 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
     1861 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
     1921 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
     1981 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
     2041 gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta caatttcctg
     2101 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatatg gtgcactctc
     2161 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     2221 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     2281 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     2341 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     2401 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     2461 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     2521 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     2581 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     2641 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     2701 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     2761 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     2821 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     2881 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     2941 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     3001 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     3061 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     3121 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     3181 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     3241 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     3301 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     3361 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     3421 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     3481 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     3541 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     3601 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     3661 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     3721 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     3781 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     3841 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     3901 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     3961 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     4021 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     4081 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     4141 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     4201 tttgct
//
