LOCUS       Exported                4285 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  E17/Q157 TAG GFP reporter.
ACCESSION   .
VERSION     .
KEYWORDS    GFP E17TAG/Q157TAG
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 13, 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
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADK*KNGIKA
                     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 tagaagaatg 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
//
