LOCUS Exported 4285 bp ds-DNA circular SYN 12-MAY-2021 DEFINITION E17TAG GFP reporter. ACCESSION . VERSION . KEYWORDS GFP E17TAG SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 4285) 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 4285) AUTHORS . TITLE Direct Submission JOURNAL Exported May 12, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..4285 /organism="synthetic DNA construct" /mol_type="other DNA" CDS complement(152..775) /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 683..702 /label=Tet-R /note="Tetracycline resistance gene, reverse primer" protein_bind 824..840 /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 847..863 /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 872..945 /label=PLtetO-1 promoter /note="modified phage lambda PL promoter with tet operator sites (Lutz and Bujard, 1997)" protein_bind 872..890 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" protein_bind 897..915 /gene="tetO" /label=tet operator /bound_moiety="tetracycline repressor TetR" /note="bacterial operator O2 for the tetR and tetA genes" RBS 955..966 /note="strong bacterial ribosome binding site (Elowitz and Leibler, 2000)" CDS 973..1686 /codon_start=1 /product="GFP variant optimized for excitation by UV light" /label=GFPuv /translation="MSKGEELFTGVVPILV*LDGDVNGHKFSVSGEGEGDATYGKLTLK FICTTGKLPVPWPTLVTTFSYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDG NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIKA NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE FVTAAGITHGMDELYK" primer_bind 1626..1646 /label=GFP-F /note="GFP, forward primer. Does NOT anneal to EGFP" CDS 1693..1710 /codon_start=1 /product="6xHis affinity tag" /label=6xHis /translation="HHHHHH" primer_bind complement(1786..1804) /label=T7 Term /note="T7 terminator, reverse primer" terminator 1800..1847 /label=T7 terminator /note="transcription terminator for bacteriophage T7 RNA polymerase" rep_origin 1944..2302 /direction=RIGHT /label=f1 ori /note="f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis" primer_bind complement(2031..2050) /label=F1ori-R /note="F1 origin, reverse primer" primer_bind 2241..2262 /label=F1ori-F /note="F1 origin, forward primer" promoter 2327..2431 /gene="bla" /label=AmpR promoter CDS 2432..3292 /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(2650..2669) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 3463..4051 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3952..3971 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 4205..4222 /label=L4440 /note="L4440 vector, forward primer" ORIGIN 1 gatctatgat atcgattttt atttgatgcc tggcagttcc ctactctcgc atggggagac 61 cccacactac catcggcgct acggcgtttc acttctgagt tcggcatggg gtcaggtggg 121 accaccgcgc tactgccgcc aggcatctag attaagaccc actttcacat ttaagttgtt 181 tttctaatcc gcatatgatc aattcaaggc cgaataagaa ggctggctct gcaccttggt 241 gatcaaataa ttcgatagct tgtcgtaata atggcggcat actatcagta gtaggtgttt 301 ccctttcttc tttagcgact tgatgctctt gatcttccaa tacgcaacct aaagtaaaat 361 gccccacagc gctgagtgca tataatgcat tctctagtga aaaaccttgt tggcataaaa 421 aggctaattg attttcgaga gtttcatact gtttttctgt aggccgtgta cctaaatgta 481 cttttgctcc atcgcgatga cttagtaaag cacatctaaa acttttagcg ttattacgta 541 aaaaatcttg ccagctttcc ccttctaaag ggcaaaagtg agtatggtgc ctatctaaca 601 tctcaatggc taaggcgtcg agcaaagccc gcttattttt tacatgccaa tacaatgtag 661 gctgctctac acctagcttc tgggcgagtt tacgggttgt taaaccttcg attccgacct 721 cattaagcag ctctaatgcg ctgttaatca ctttactttt atctaatctg gacatgtcga 781 atatagacgt cggtcagtgc gtcctgctga tgtgctcagt atcttgttat ccgctcacaa 841 tgtcaattgt tatccgctca caattctcga gtccctatca gtgatagaga ttgacatccc 901 tatcagtgat agagatactg agcacatcag caggacgcac tgaccgaatt cattaaagag 961 gagaaaggta ccatgagtaa aggagaagaa cttttcactg gagttgtccc aattcttgtt 1021 tagttagatg gtgatgttaa tgggcacaaa ttttctgtca gtggagaggg tgaaggtgat 1081 gcaacatacg gaaaacttac ccttaaattt atttgcacta ctggaaaact acctgttcca 1141 tggccaacac ttgtcactac tttctcttat ggtgttcaat gcttttcccg ttatccggat 1201 cacatgaaac ggcatgactt tttcaagagt gccatgcccg aaggttatgt acaggaacgc 1261 actatatctt tcaaagatga cgggaactac aagacgcgtg ctgaagtcaa gtttgaaggt 1321 gatacccttg ttaatcgtat cgagttaaaa ggtattgatt ttaaagaaga tggaaacatt 1381 ctcggacaca aactcgaata caactataac tcacacaatg tatacatcac ggcagacaaa 1441 caaaagaatg gaatcaaagc taacttcaaa attcgccaca acattgaaga tggatccgtt 1501 caactagcag accattatca acaaaatact ccaattggcg atggccctgt ccttttacca 1561 gacaaccatt acctgtcgac acaatctgcc ctttcgaaag atcccaacga aaagcgtgac 1621 cacatggtcc ttcttgagtt tgtaactgct gctgggatta cacatggcat ggatgagctc 1681 tacaaactcg agcaccacca ccaccaccac tgaagggcga gctcaattca aagcttgatc 1741 cggctgctaa caaagcccga aaggaagctg agttggctgc tgccaccgct gagcaataac 1801 tagcataacc ccttggggcc tctaaacggg tcttgagggg ttttttgctg aaaggaggaa 1861 ctatatccgg atctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt 1921 gcgcagcctg aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt 1981 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc 2041 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg 2101 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta 2161 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt 2221 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat 2281 ctcggtctat tcttttgatt taggtggcac ttttcgggga aatgtgcgcg gaacccctat 2341 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata 2401 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct 2461 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa 2521 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa 2581 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt 2641 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg 2701 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca 2761 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa 2821 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt 2881 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc 2941 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa 3001 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga 3061 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc 3121 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga 3181 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga 3241 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga 3301 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat 3361 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt 3421 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct 3481 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc 3541 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc 3601 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc 3661 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc 3721 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg 3781 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata 3841 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta 3901 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc 3961 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg 4021 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt 4081 cctggccttt tgctggcctt ttgctcacat gttctttcct gcgttatccc ctgattctgt 4141 ggataaccgt attaccgcct ttgagtgagc tgataccgct cgccgcagcc gaacgaccga 4201 gcgcagcgag tcagtgagcg aggaagcgga agagcgccca atacgcaaac cgcctctccc 4261 cgcgcgttgg ccgattcatt aatgc //