LOCUS       sequence-440999-        4348 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  S2TAG 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 S2TAG
FEATURES             Location/Qualifiers
     source          1..4348
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    35..51
                     /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    58..74
                     /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(83..101,102..107,108..124,125..130,131..156)
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator 
                     sites (Lutz and Bujard, 1997)"
     protein_bind    83..101
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    108..126
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             166..177
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             184..897
                     /codon_start=1
                     /product="GFP variant optimized for excitation by UV light"
                     /label=GFPuv
                     /translation="M*KGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     primer_bind     complement(214..242)
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     primer_bind     837..857
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     CDS             904..921
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(997..1015)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      1011..1058
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      1155..1610
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1242..1261)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1452..1473
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1636..1740
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1741..1809,1810..2601)
                     /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(1959..1978)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2772..3360
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3261..3280
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3514..3531
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             complement(3711..4334)
                     /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     4242..4261
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
ORIGIN
        1 tcggtcagtg cgtcctgctg atgtgctcag tatcttgtta tccgctcaca atgtcaattg
       61 ttatccgctc acaattctcg agtccctatc agtgatagag attgacatcc ctatcagtga
      121 tagagatact gagcacatca gcaggacgca ctgaccgaat tcattaaaga ggagaaaggt
      181 accatgtaga aaggagaaga acttttcact ggagttgtcc caattcttgt tgaattagat
      241 ggtgatgtta atgggcacaa attttctgtc agtggagagg gtgaaggtga tgcaacatac
      301 ggaaaactta cccttaaatt tatttgcact actggaaaac tacctgttcc atggccaaca
      361 cttgtcacta ctttctctta tggtgttcaa tgcttttccc gttatccgga tcacatgaaa
      421 cggcatgact ttttcaagag tgccatgccc gaaggttatg tacaggaacg cactatatct
      481 ttcaaagatg acgggaacta caagacgcgt gctgaagtca agtttgaagg tgataccctt
      541 gttaatcgta tcgagttaaa aggtattgat tttaaagaag atggaaacat tctcggacac
      601 aaactcgaat acaactataa ctcacacaat gtatacatca cggcagacaa acaaaagaat
      661 ggaatcaaag ctaacttcaa aattcgccac aacattgaag atggatccgt tcaactagca
      721 gaccattatc aacaaaatac tccaattggc gatggccctg tccttttacc agacaaccat
      781 tacctgtcga cacaatctgc cctttcgaaa gatcccaacg aaaagcgtga ccacatggtc
      841 cttcttgagt ttgtaactgc tgctgggatt acacatggca tggatgagct ctacaaactc
      901 gagcaccacc accaccacca ctgaagggcg agctcaattc aaagcttgat ccggctgcta
      961 acaaagcccg aaaggaagct gagttggctg ctgccaccgc tgagcaataa ctagcataac
     1021 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaaggagga actatatccg
     1081 gatctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
     1141 gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1201 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1261 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1321 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1381 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1441 gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta tctcggtcta
     1501 ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa atgagctgat
     1561 ttaacaaaaa tttaacgcga attttaacaa aatattaacg cttacaattt aggtggcact
     1621 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     1681 tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt
     1741 atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct
     1801 gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca
     1861 cgagtgggtt acatcgaact ggatctcaac agcggtaaga tccttgagag ttttcgcccc
     1921 gaagaacgtt ttccaatgat gagcactttt aaagttctgc tatgtggcgc ggtattatcc
     1981 cgtattgacg ccgggcaaga gcaactcggt cgccgcatac actattctca gaatgacttg
     2041 gttgagtact caccagtcac agaaaagcat cttacggatg gcatgacagt aagagaatta
     2101 tgcagtgctg ccataaccat gagtgataac actgcggcca acttacttct gacaacgatc
     2161 ggaggaccga aggagctaac cgcttttttg cacaacatgg gggatcatgt aactcgcctt
     2221 gatcgttggg aaccggagct gaatgaagcc ataccaaacg acgagcgtga caccacgatg
     2281 cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg gcgaactact tactctagct
     2341 tcccggcaac aattaataga ctggatggag gcggataaag ttgcaggacc acttctgcgc
     2401 tcggcccttc cggctggctg gtttattgct gataaatctg gagccggtga gcgtgggtct
     2461 cgcggtatca ttgcagcact ggggccagat ggtaagccct cccgtatcgt agttatctac
     2521 acgacgggga gtcaggcaac tatggatgaa cgaaatagac agatcgctga gataggtgcc
     2581 tcactgatta agcattggta actgtcagac caagtttact catatatact ttagattgat
     2641 ttaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     2701 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     2761 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     2821 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     2881 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     2941 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3001 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3061 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3121 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3181 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3241 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3301 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3361 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3421 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     3481 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagccaaa
     3541 gggcccaagg cccagtcttt cgactgagcc tttcgtttta tttgatgcct ggcagttccc
     3601 tactctcgca tggggagacc ccacactacc atcggcgcta cggcgtttca cttctgagtt
     3661 cggcatgggg tcaggtggga ccaccgcgct actgccgcca ggcatctaga ttaagaccca
     3721 ctttcacatt taagttgttt ttctaatccg catatgatca attcaaggcc gaataagaag
     3781 gctggctctg caccttggtg atcaaataat tcgatagctt gtcgtaataa tggcggcata
     3841 ctatcagtag taggtgtttc cctttcttct ttagcgactt gatgctcttg atcttccaat
     3901 acgcaaccta aagtaaaatg ccccacagcg ctgagtgcat ataatgcatt ctctagtgaa
     3961 aaaccttgtt ggcataaaaa ggctaattga ttttcgagag tttcatactg tttttctgta
     4021 ggccgtgtac ctaaatgtac ttttgctcca tcgcgatgac ttagtaaagc acatctaaaa
     4081 cttttagcgt tattacgtaa aaaatcttgc cagctttccc cttctaaagg gcaaaagtga
     4141 gtatggtgcc tatctaacat ctcaatggct aaggcgtcga gcaaagcccg cttatttttt
     4201 acatgccaat acaatgtagg ctgctctaca cctagcttct gggcgagttt acgggttgtt
     4261 aaaccttcga ttccgacctc attaagcagc tctaatgcgc tgttaatcac tttactttta
     4321 tctaatctgg acatgtcgaa tatagacg
//
