LOCUS       sequence-441002-        4348 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  E17/Q157 TAG 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/Q157TAG
FEATURES             Location/Qualifiers
     source          1..4348
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    37..53
                     /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    60..76
                     /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(85..103,104..109,110..126,127..132,133..158)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    85..103
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    110..128
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             168..179
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             186..899
                     /codon_start=1
                     /product="GFP variant optimized for excitation by UV light"
                     /label=GFPuv
                     /translation="MSKGEELFTGVVPILV*LDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADK*KNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     839..859
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             906..923
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(999..1017)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1013..1060
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1157..1612
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1244..1263)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1454..1475
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1638..1742
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1743..1811,1812..2603)
                     /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(1961..1980)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2774..3362
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3263..3282
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3516..3533
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3713..4336)
                     /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     4244..4263
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
ORIGIN
        1 cgtcggtcag tgcgtcctgc tgatgtgctc agtatcttgt tatccgctca caatgtcaat
       61 tgttatccgc tcacaattct cgagtcccta tcagtgatag agattgacat ccctatcagt
      121 gatagagata ctgagcacat cagcaggacg cactgaccga attcattaaa gaggagaaag
      181 gtaccatgag taaaggagaa gaacttttca ctggagttgt cccaattctt gtttagttag
      241 atggtgatgt taatgggcac aaattttctg tcagtggaga gggtgaaggt gatgcaacat
      301 acggaaaact tacccttaaa tttatttgca ctactggaaa actacctgtt ccatggccaa
      361 cacttgtcac tactttctct tatggtgttc aatgcttttc ccgttatccg gatcacatga
      421 aacggcatga ctttttcaag agtgccatgc ccgaaggtta tgtacaggaa cgcactatat
      481 ctttcaaaga tgacgggaac tacaagacgc gtgctgaagt caagtttgaa ggtgataccc
      541 ttgttaatcg tatcgagtta aaaggtattg attttaaaga agatggaaac attctcggac
      601 acaaactcga atacaactat aactcacaca atgtatacat cacggcagac aaatagaaga
      661 atggaatcaa agctaacttc aaaattcgcc acaacattga agatggatcc gttcaactag
      721 cagaccatta tcaacaaaat actccaattg gcgatggccc tgtcctttta ccagacaacc
      781 attacctgtc gacacaatct gccctttcga aagatcccaa cgaaaagcgt gaccacatgg
      841 tccttcttga gtttgtaact gctgctggga ttacacatgg catggatgag ctctacaaac
      901 tcgagcacca ccaccaccac cactgaaggg cgagctcaat tcaaagcttg atccggctgc
      961 taacaaagcc cgaaaggaag ctgagttggc tgctgccacc gctgagcaat aactagcata
     1021 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaaggag gaactatatc
     1081 cggatctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc
     1141 ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt
     1201 acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc
     1261 ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct
     1321 ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga ttagggtgat
     1381 ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc
     1441 acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc tatctcggtc
     1501 tattcttttg atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg
     1561 atttaacaaa aatttaacgc gaattttaac aaaatattaa cgcttacaat ttaggtggca
     1621 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     1681 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     1741 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     1801 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     1861 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     1921 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     1981 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     2041 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     2101 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     2161 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     2221 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     2281 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     2341 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     2401 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     2461 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     2521 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     2581 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     2641 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     2701 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     2761 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     2821 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     2881 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     2941 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3001 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     3061 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3121 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     3181 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     3241 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     3301 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     3361 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     3421 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     3481 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcca
     3541 aagggcccaa ggcccagtct ttcgactgag cctttcgttt tatttgatgc ctggcagttc
     3601 cctactctcg catggggaga ccccacacta ccatcggcgc tacggcgttt cacttctgag
     3661 ttcggcatgg ggtcaggtgg gaccaccgcg ctactgccgc caggcatcta gattaagacc
     3721 cactttcaca tttaagttgt ttttctaatc cgcatatgat caattcaagg ccgaataaga
     3781 aggctggctc tgcaccttgg tgatcaaata attcgatagc ttgtcgtaat aatggcggca
     3841 tactatcagt agtaggtgtt tccctttctt ctttagcgac ttgatgctct tgatcttcca
     3901 atacgcaacc taaagtaaaa tgccccacag cgctgagtgc atataatgca ttctctagtg
     3961 aaaaaccttg ttggcataaa aaggctaatt gattttcgag agtttcatac tgtttttctg
     4021 taggccgtgt acctaaatgt acttttgctc catcgcgatg acttagtaaa gcacatctaa
     4081 aacttttagc gttattacgt aaaaaatctt gccagctttc cccttctaaa gggcaaaagt
     4141 gagtatggtg cctatctaac atctcaatgg ctaaggcgtc gagcaaagcc cgcttatttt
     4201 ttacatgcca atacaatgta ggctgctcta cacctagctt ctgggcgagt ttacgggttg
     4261 ttaaaccttc gattccgacc tcattaagca gctctaatgc gctgttaatc actttacttt
     4321 tatctaatct ggacatgtcg aatataga
//
