LOCUS       sequence-339028-        3687 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  QUAS directly upstream of a T7 promoter with a TetO operator. 
            Expression of TetR in BL21(DE3) E. coli. GFP is expressed according 
            to presence of aTc. Contains the ColE1 origin of replication..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3687)
  AUTHORS   MacDonald IC, Seamons TR, Emmons JC, Javdan SB, Deans TL
  TITLE     Enhanced regulation of prokaryotic gene expression by a eukaryotic 
            transcriptional activator.
  JOURNAL   Nat Commun. 2021 Jul 5;12(1):4109. doi: 10.1038/s41467-021-24434-9.
  PUBMED    34226549
REFERENCE   2  (bases 1 to 3687)
  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: "2021-07-5"; volume: "12(1):4109. doi"; pages: " 
            10.1038/s41467-021-24434-9"
COMMENT     Sequence Label: QUAS-0-T7-TetO-GFP_T7-LacO-TetR_ColE1
FEATURES             Location/Qualifiers
     source          1..3687
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(join(1..792,793..861))
                     /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     624..643
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(862..966)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        1002..1020
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1021..1039
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             join(1067..1069,1070..1072,1073..1783)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(1067..1069,1070..1072,1073..1783)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1112..1133)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1373..1392)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1720..1741
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      1856..1903
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        1948..1966
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1967..1991
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     RBS             2007..2027
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             2036..2656
                     /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(2109..2128)
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     terminator      2699..2746
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      complement(2906..3494)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2986..3005)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     terminator      complement(3582..3676)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
ORIGIN
        1 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
       61 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
      121 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
      181 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
      241 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
      301 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
      361 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
      421 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
      481 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
      541 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
      601 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
      661 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
      721 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
      781 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
      841 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
      901 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
      961 tccgcgcgct agcggtaccg ctatagggta atcgcttatc ctaatacgac tcactatagg
     1021 tccctatcag tgatagagag tttaacttta agaaggagga gtacatatgg tgagcaaggg
     1081 cgaggagctg ttcaccgggg tggtgcccat cctggtcgag ctggacggcg acgtaaacgg
     1141 ccacaagttc agcgtgtccg gcgagggcga gggcgatgcc acctacggca agctgaccct
     1201 gaagttcatc tgcaccaccg gcaagctgcc cgtgccctgg cccaccctcg tgaccaccct
     1261 gacctacggc gtgcagtgct tcagccgcta ccccgaccac atgaagcagc acgacttctt
     1321 caagtccgcc atgcccgaag gctacgtcca ggagcgcacc atcttcttca aggacgacgg
     1381 caactacaag acccgcgccg aggtgaagtt cgagggcgac accctggtga accgcatcga
     1441 gctgaagggc atcgacttca aggaggacgg caacatcctg gggcacaagc tggagtacaa
     1501 ctacaacagc cacaacgtct atatcatggc cgacaagcag aagaacggca tcaaggtgaa
     1561 cttcaagatc cgccacaaca tcgaggacgg cagcgtgcag ctcgccgacc actaccagca
     1621 gaacaccccc atcggcgacg gccccgtgct gctgcccgac aaccactacc tgagcaccca
     1681 gtccgccctg agcaaagacc ccaacgagaa gcgcgatcac atggtcctgc tggagttcgt
     1741 gaccgccgcc gggatcactc tcggcatgga cgagctgtac aagtccggac tcagatctcg
     1801 agcggcgaac gacgaaaact actctgaaaa ctacgcggac gcgtcttaga agcttctagc
     1861 ataacccctt ggggcctcta aacgggtctt gaggggtttt ttgcccgggc tcagggcgtt
     1921 gaattccgct agccgatccc gcgaaattaa tacgactcac tataggggaa ttgtgagcgg
     1981 ataacaattc ccgactagaa ataattttgt ttaactttaa gaaggaggag agtccatgtc
     2041 tagattagat aaaagtaaag tgattaacag cgcattagag ctgcttaatg aggtcggaat
     2101 cgaaggttta acaacccgta aactcgccca gaagctaggt gtagagcagc ctacattgta
     2161 ttggcatgta aaaaataagc gggctttgct cgacgcctta gccattgaga tgttagatag
     2221 gcaccatact cacttttgcc ctttagaagg ggaaagctgg caagattttt tacgtaataa
     2281 cgctaaaagt tttagatgtg ctttactaag tcatcgcgat ggagcaaaag tacatttagg
     2341 tacacggcct acagaaaaac agtatgaaac tctcgaaaat caattagcct ttttatgcca
     2401 acaaggtttt tcactagaga atgcattata tgcactcagc gctgtggggc attttacttt
     2461 aggttgcgta ttggaagatc aagagcatca agtcgctaaa gaagaaaggg aaacacctac
     2521 tactgatagt atgccgccat tattacgaca agctatcgaa ttatttgatc accaaggtgc
     2581 agagccagcc ttcttattcg gccttgaatt gatcatatgc ggattagaaa aacaacttaa
     2641 atgtgaaagt gggtccgcgt acagcggatc ccgggaattc agatcttatt aaaagcttct
     2701 agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttggcgg ccgccctagg
     2761 gcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     2821 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     2881 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     2941 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3001 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3061 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     3121 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     3181 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     3241 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     3301 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta
     3361 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     3421 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     3481 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     3541 aaaactcacg ttaagggatt ttggtcatga ctagtgcttg gattctcacc aataaaaaac
     3601 gcccggcggc aaccgagcgt tctgaacaaa tccagatgga gttctgaggt cattactgga
     3661 tctatcaaca ggagtccaag cgagctc
//
