LOCUS       62056_20315        3064 bp DNA     circular SYN 19-APR-2020
DEFINITION  Cloning vector pSyn_Mz_Mj_Eser_sfGFP_151_190-200(dest), complete 
            sequence.
ACCESSION   MN882188
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3064)
  AUTHORS   Fischer EC, Hashimoto K, Zhang Y, Feldman AW, Dien VT, Karadeema RJ,
            Adhikary R, Ledbetter MP, Krishnamurthy R, Romesberg FE.
  TITLE     New codons for efficient production of unnatural proteins in a 
            semi-synthetic organism
  JOURNAL   Nat. Chem. Biol. (2020) In press
REFERENCE   2  (bases 1 to 3064)
  AUTHORS   Fischer EC.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-DEC-2019) Department of Chemistry, The Scripps 
            Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 
            92037, USA
REFERENCE   3  (bases 1 to 3064)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. Chem. 
            Biol. (2020) In press"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (27-DEC-2019) Department of Chemistry, The Scripps Research 
            Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, USA"
FEATURES             Location/Qualifiers
     source          1..3064
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      48..593
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin."
     promoter        746..764
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    766..784
                     /label=tet operator
                     /note="bacterial operator O2 for the tetR and tetA genes"
     RBS             799..821
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     misc_feature    1248..1265
                     /label=Y151 Golden Gate destination site
                     /note="Y151 Golden Gate destination site"
     misc_feature    1339..1356
                     /label=D190-Y200 Golden Gate destination site
                     /note="D190-Y200 Golden Gate destination site"
     CDS             1449..1538
                     /label=Twin-Strep-tag
                     /note="two Strep-Tag(R)II moieties connected by a linker 
                     for enhanced binding to Strep-Tactin(R), an engineered form
                     of streptavidin (Schmidt et al., 2013)"
     terminator      1589..1636
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     regulatory      1666..1684
                     /note="promoter for bacteriophage T7 RNA polymerase; T7 
                     promoter"
                     /regulatory_class="promoter"
     protein_bind    1685..1709
                     /label=lac operator
                     /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)."
     misc_feature    1721..1725
                     /label=5' proK leader
                     /note="5' proK leader"
     misc_feature    1726..1773
                     /note="tRNA-Pyl; Methanosarcina mazei tRNAPyl(dest)"
     misc_feature    1733..1750
                     /label=tRNAPyl Golden Gate destination site
                     /note="tRNAPyl Golden Gate destination site"
     misc_feature    1774..1812
                     /label=3' proK trailer
                     /note="3' proK trailer"
     regulatory      1813..1860
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase; T7 terminator"
                     /regulatory_class="terminator"
     regulatory      1907..1925
                     /note="promoter for bacteriophage T7 RNA polymerase; T7 
                     promoter"
                     /regulatory_class="promoter"
     protein_bind    1926..1950
                     /label=lacI binding site
                     /bound_moiety="lacI"
                     /note="lac repressor binds to the lac operator to inhibit 
                     transcription in E. coli; lac operator"
     misc_feature    1962..1966
                     /label=5' proK leader
                     /note="5' proK leader"
     misc_feature    1967..2009
                     /note="tRNA; Methanocaldococcus jannaschii tRNApAzF(dest)"
     misc_feature    1977..1994
                     /label=tRNApAzF Golden Gate destination site
                     /note="tRNApAzF Golden Gate destination site"
     misc_feature    2010..2048
                     /label=3' proK trailer
                     /note="3' proK trailer"
     regulatory      2049..2096
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase; T7 terminator"
                     /regulatory_class="terminator"
     regulatory      2181..2199
                     /note="promoter for bacteriophage T7 RNA polymerase; T7 
                     promoter"
                     /regulatory_class="promoter"
     protein_bind    2200..2217
                     /label=tetracycline repressor TetR binding site
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes;
                     tet operator"
     misc_feature    2230..2236
                     /label=5' serT leader
                     /note="5' serT leader"
     misc_feature    2237..2298
                     /label=Escherichia coli tRNASer(dest)
                     /note="Escherichia coli tRNASer(dest)"
     misc_feature    2251..2268
                     /label=tRNASer Golden Gate destination site
                     /note="tRNASer Golden Gate destination site"
     misc_feature    2299..2341
                     /label=3' serT trailer
                     /note="3' serT trailer"
     regulatory      2342..2389
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase; T7 terminator"
                     /regulatory_class="terminator"
     promoter        2448..2495
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     CDS             2514..2885
                     /label=BleoR
                     /note="antibiotic-binding protein"
     terminator      2900..2994
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
ORIGIN
        1 tgcactgaaa tctagtaata ttttatctga ttaataagat gatcttcttg agatcgtttt
       61 ggtctgcgcg taatctcttg ctctgaaaac gaaaaaaccg ccttgcaggg cggtttttcg
      121 aaggttctct gagctaccaa ctctttgaac cgaggtaact ggcttggagg agcgcagtca
      181 ccaaaacttg tcctttcagt ttagccttaa ccggcgcatg acttcaagac taactcctct
      241 aaatcaatta ccagtggctg ctgccagtgg tgcttttgca tgtctttccg ggttggactc
      301 aagacgatag ttaccggata aggcgcagcg gtcggactga acggggggtt cgtgcataca
      361 gtccagcttg gagcgaactg cctacccgga actgagtgtc aggcgtggaa tgagacaaac
      421 gcggccataa cagcggaatg acaccggtaa accgaaaggc aggaacagga gagcgcacga
      481 gggagccgcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccaccact
      541 gatttgagcg tcagatttcg tgatgcttgt caggggggcg gagcctatgg aaaaacggct
      601 ttgccgcggc cctctcactt ccctgttaag tatcttcctg gcatcttcca ggaaatctcc
      661 gccccgttcg taagccattt ccgctcgccg cagtcgaacg accgagcgta gcgagtcagt
      721 gagcgaggaa gcggaatata tcccttaata cgactcacta tagggtccct atcagtgata
      781 gagaggtcta gaaataattt tgtttaactt taagaaggag atatacatat gtcgaaaggc
      841 gaagaactgt ttacgggagt ggtgcctatc ctggtagagc tcgacggaga tgtaaacggt
      901 cacaaatttt cagtccgcgg ggaaggcgaa ggcgatgcga ccaacggtaa attaactttg
      961 aagtttattt gcaccaccgg caaattaccg gtgccttggc cgacgcttgt gacgaccctg
     1021 acttacgggg tgcagtgttt cagtcgctac ccagatcaca tgaaacgcca tgacttcttc
     1081 aaatctgcga tgccggaagg ctatgtgcag gaacgtacaa ttagctttaa agacgacggc
     1141 acgtataaaa cgcgggcaga ggttaaattt gagggagata ccctggtaaa ccgtattgaa
     1201 ctgaaaggca tcgattttaa agaagatggg aacatcttgg gccacaagag accggtaccg
     1261 gtctcggaat caaagcaaat ttcaagatcc gtcataacgt ggaggacggt tccgtgcagc
     1321 ttgcagatca ctatcagaga gaccggtacc ggtctcaaga cccgaacgag aagcgtgatc
     1381 acatggtgct gcttgaattt gttaccgcgg caggtattac acacggcatg gatgagttgt
     1441 ataagggatc cgcttggagc cacccgcagt tcgagaaagg tggaggttcc ggaggtggat
     1501 cgggaggttc ggcgtggagc cacccgcagt tcgaaaaata aaagcttaat tagctgagct
     1561 tggactccct gccaccgctg agcaataact agcataaccc cttggggcct ctaaacgggt
     1621 cttgaggggt tttttgctga aaggaggaac tatatccgga ttggttaata cgactcacta
     1681 taggggaatt gtgagcggat aacaattccc ctctagaaaa gcattggaaa ccgagaccgg
     1741 taccggtctc ttagattccc ggggtttccg ccaaattcga aaagcctgct caacgagcag
     1801 gcttttttgc atctagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     1861 cctgaacgag caggctgctg aaaggaggaa ctatatccgg attggttaat acgactcact
     1921 ataggggaat tgtgagcgga taacaattcc cctctagaaa agcattccgg cggtaggaga
     1981 ccggtaccgg tctcccctcc gccggaccaa attcgaaaag cctgctcaac gagcaggctt
     2041 ttttgcatct agcataaccc cttggggcct ctaaacgggt cttgaggggt tttttgcctg
     2101 aacgagcagg cttttttgca taagcttcct agtggcagcg gctaactaat gctgaaagga
     2161 ggaactatat ccggattggt taatacgact cactataggt ccctatcagt gatagagtcc
     2221 tctagaaaat cattccggaa gtgtggccga gagaccggta ccggtctctc cagagttcga
     2281 atctctgcgc ttccgccaaa taagataagg ggttagctaa atgctaaccc ctttttcttt
     2341 tctagcataa ccccttgggg cctctaaacg ggtcttgagg ggttttttgc ctgaacgagc
     2401 aggctttgtg cgcggaaccc ctatttgttt agattgcgga tcccgtcgtt gacaattaat
     2461 catcggcata gtatatcggc atagtataat acgacaaggt gaggaactaa accatggcca
     2521 agttgaccag tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct
     2581 ggaccgaccg gctcgggttc tcccgcgact tcgtggagga cgacttcgcc ggtgtggtcc
     2641 gggacgacgt gaccctgttc atcagcgcgg tccaggacca ggtggtgccg gacaacaccc
     2701 tggcctgggt gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg gaggtcgtgt
     2761 ccacgaactt ccgggacgcc tccgggccgg ccatgaccga gatcggcgag cagccgtggg
     2821 ggcgggagtt cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg gccgaggagc
     2881 aggactgaga gctcgcttgg actcctgttg atagatccag taatgacctc agaactccat
     2941 ctggatttgt tcagaacgct cggttgccgc cgggcgtttt ttattggtga gaatccaagc
     3001 actagctagt aacaacttat atcgtatggg gctgacttca ggtgctacat ttgaagagat
     3061 aaat
//
