LOCUS       sequence-440998-        4348 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E17TAG GFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4348)
  AUTHORS   Pirman NL, Barber KW, Aerni HR, Ma NJ, Haimovich AD, Rogulina S, 
            Isaacs FJ, Rinehart J
  TITLE     A flexible codon in genomically recoded Escherichia coli permits 
            programmable protein phosphorylation.
  JOURNAL   Nat Commun. 2015 Sep 9;6:8130. doi: 10.1038/ncomms9130.
  PUBMED    26350500
REFERENCE   2  (bases 1 to 4348)
  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: "2015-09-9"; volume: "6:8130. doi"; pages: " 
            10.1038/ncomms9130"
COMMENT     Sequence Label: GFP E17TAG
FEATURES             Location/Qualifiers
     source          1..4348
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    21..37
                     /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)."
     protein_bind    44..60
                     /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)."
     promoter        join(69..87,88..93,94..110,111..116,117..142)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    69..87
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    94..112
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             152..163
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             170..883
                     /codon_start=1
                     /product="GFP variant optimized for excitation by UV light"
                     /label=GFPuv
                     /translation="MSKGEELFTGVVPILV*LDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     823..843
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             890..907
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(983..1001)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      997..1044
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1141..1596
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1228..1247)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1438..1459
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1622..1726
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1727..1795,1796..2587)
                     /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(1945..1964)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2758..3346
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3247..3266
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3500..3517
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3697..4320)
                     /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     4228..4247
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
ORIGIN
        1 ctgctgatgt gctcagtatc ttgttatccg ctcacaatgt caattgttat ccgctcacaa
       61 ttctcgagtc cctatcagtg atagagattg acatccctat cagtgataga gatactgagc
      121 acatcagcag gacgcactga ccgaattcat taaagaggag aaaggtacca tgagtaaagg
      181 agaagaactt ttcactggag ttgtcccaat tcttgtttag ttagatggtg atgttaatgg
      241 gcacaaattt tctgtcagtg gagagggtga aggtgatgca acatacggaa aacttaccct
      301 taaatttatt tgcactactg gaaaactacc tgttccatgg ccaacacttg tcactacttt
      361 ctcttatggt gttcaatgct tttcccgtta tccggatcac atgaaacggc atgacttttt
      421 caagagtgcc atgcccgaag gttatgtaca ggaacgcact atatctttca aagatgacgg
      481 gaactacaag acgcgtgctg aagtcaagtt tgaaggtgat acccttgtta atcgtatcga
      541 gttaaaaggt attgatttta aagaagatgg aaacattctc ggacacaaac tcgaatacaa
      601 ctataactca cacaatgtat acatcacggc agacaaacaa aagaatggaa tcaaagctaa
      661 cttcaaaatt cgccacaaca ttgaagatgg atccgttcaa ctagcagacc attatcaaca
      721 aaatactcca attggcgatg gccctgtcct tttaccagac aaccattacc tgtcgacaca
      781 atctgccctt tcgaaagatc ccaacgaaaa gcgtgaccac atggtccttc ttgagtttgt
      841 aactgctgct gggattacac atggcatgga tgagctctac aaactcgagc accaccacca
      901 ccaccactga agggcgagct caattcaaag cttgatccgg ctgctaacaa agcccgaaag
      961 gaagctgagt tggctgctgc caccgctgag caataactag cataacccct tggggcctct
     1021 aaacgggtct tgaggggttt tttgctgaaa ggaggaacta tatccggatc tggcgtaata
     1081 gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat ggcgaatggg
     1141 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     1201 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     1261 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     1321 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     1381 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     1441 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     1501 aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta
     1561 acgcgaattt taacaaaata ttaacgctta caatttaggt ggcacttttc ggggaaatgt
     1621 gcgcggaacc cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag
     1681 acaataaccc tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca
     1741 tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc
     1801 agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat
     1861 cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc
     1921 aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg
     1981 gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc
     2041 agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat
     2101 aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga
     2161 gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc
     2221 ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc
     2281 aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt
     2341 aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc
     2401 tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc
     2461 agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca
     2521 ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca
     2581 ttggtaactg tcagaccaag tttactcata tatactttag attgatttaa aacttcattt
     2641 ttaatttaaa aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta
     2701 acgtgagttt tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg
     2761 agatcctttt tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc
     2821 ggtggtttgt ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag
     2881 cagagcgcag ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa
     2941 gaactctgta gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc
     3001 cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc
     3061 gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta
     3121 caccgaactg agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag
     3181 aaaggcggac aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct
     3241 tccaggggga aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga
     3301 gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc
     3361 ggccttttta cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt
     3421 atcccctgat tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg
     3481 cagccgaacg accgagcgca gcgagtcagt gagcgaggaa gccaaagggc ccaaggccca
     3541 gtctttcgac tgagcctttc gttttatttg atgcctggca gttccctact ctcgcatggg
     3601 gagaccccac actaccatcg gcgctacggc gtttcacttc tgagttcggc atggggtcag
     3661 gtgggaccac cgcgctactg ccgccaggca tctagattaa gacccacttt cacatttaag
     3721 ttgtttttct aatccgcata tgatcaattc aaggccgaat aagaaggctg gctctgcacc
     3781 ttggtgatca aataattcga tagcttgtcg taataatggc ggcatactat cagtagtagg
     3841 tgtttccctt tcttctttag cgacttgatg ctcttgatct tccaatacgc aacctaaagt
     3901 aaaatgcccc acagcgctga gtgcatataa tgcattctct agtgaaaaac cttgttggca
     3961 taaaaaggct aattgatttt cgagagtttc atactgtttt tctgtaggcc gtgtacctaa
     4021 atgtactttt gctccatcgc gatgacttag taaagcacat ctaaaacttt tagcgttatt
     4081 acgtaaaaaa tcttgccagc tttccccttc taaagggcaa aagtgagtat ggtgcctatc
     4141 taacatctca atggctaagg cgtcgagcaa agcccgctta ttttttacat gccaatacaa
     4201 tgtaggctgc tctacaccta gcttctgggc gagtttacgg gttgttaaac cttcgattcc
     4261 gacctcatta agcagctcta atgcgctgtt aatcacttta cttttatcta atctggacat
     4321 gtcgaatata gacgtcggtc agtgcgtc
//
