LOCUS       62056_20330        3120 bp DNA     circular SYN 19-APR-2020
DEFINITION  Cloning vector pSyn_Mz_sfGFP_190(dest), complete sequence.
ACCESSION   MN882185
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3120)
  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 3120)
  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 3120)
  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..3120
                     /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    1379..1396
                     /label=D190 Golden Gate destination site
                     /note="D190 Golden Gate destination site"
     CDS             1531..1554
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
     CDS             1591..1614
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
     terminator      1665..1712
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     regulatory      1742..1760
                     /note="promoter for bacteriophage T7 RNA polymerase; T7 
                     promoter"
                     /regulatory_class="promoter"
     protein_bind    1761..1785
                     /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    1797..1801
                     /label=5' proK leader
                     /note="5' proK leader"
     misc_feature    1802..1849
                     /note="tRNA-Pyl; Methanosarcina mazei tRNAPyl(dest)"
     misc_feature    1809..1826
                     /label=tRNAPyl Golden Gate destination site
                     /note="tRNAPyl Golden Gate destination site"
     misc_feature    1850..1888
                     /label=3' proK trailer
                     /note="3' proK trailer"
     regulatory      1889..1936
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase; T7 terminator"
                     /regulatory_class="terminator"
     regulatory      1974..2080
                     /label=proK promoter
                     /note="proK promoter"
                     /regulatory_class="promoter"
     misc_RNA        2101..2176
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     regulatory      2184..2219
                     /label=proK terminator
                     /note="proK terminator"
                     /regulatory_class="terminator"
     regulatory      2232..2337
                     /label=proK promoter
                     /note="proK promoter"
                     /regulatory_class="promoter"
     misc_RNA        2356..2427
                     /note="guide RNA scaffold for the CRISPR/Cas9 system; gRNA 
                     scaffold"
     regulatory      2435..2470
                     /label=proK terminator
                     /note="proK terminator"
                     /regulatory_class="terminator"
     promoter        2504..2551
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     CDS             2570..2941
                     /label=BleoR
                     /note="antibiotic-binding protein"
     terminator      2956..3050
                     /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 gccacaaact cgagtacaac
     1261 tttaactcac acaatgtata catcacggca gacaaacaaa agaatggaat caaagcaaat
     1321 ttcaagatcc gtcataacgt ggaggacggt tccgtgcagc ttgcagatca ctatcagaga
     1381 gaccggtacc ggtctcataa ccactacctc tctactcaga gtgttttatc gaaagacccg
     1441 aacgagaagc gtgatcacat ggtgctgctt gaatttgtta ccgcggcagg tattacacac
     1501 ggcatggatg agttgtataa gggatccgct tggagccacc cgcagttcga gaaaggtgga
     1561 ggttccggag gtggatcggg aggttcggcg tggagccacc cgcagttcga aaaataaaag
     1621 cttaattagc tgagcttgga ctccctgcca ccgctgagca ataactagca taaccccttg
     1681 gggcctctaa acgggtcttg aggggttttt tgctgaaagg aggaactata tccggattgg
     1741 ttaatacgac tcactatagg ggaattgtga gcggataaca attcccctct agaaaagcat
     1801 tggaaaccga gaccggtacc ggtctcttag attcccgggg tttccgccaa attcgaaaag
     1861 cctgctcaac gagcaggctt ttttgcatct agcataaccc cttggggcct ctaaacgggt
     1921 cttgaggggt tttttgcctg aacgagcagg cttttttgca taagcttcct agtggcagcg
     1981 gctaactaag cggcctgctg actttctcgc cgatcaaaag gcattttgct attaagggat
     2041 tgacgagggc gtatctgcgc agtaagatgc gccccgcatt ggagacgcca tggcgtctcg
     2101 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
     2161 ggcaccgagt cggtgctttt tttaattcga aaagcctgct caacgagcag gcttttttgg
     2221 tcgacagtag tggcagcggc taactaagcg gcctgctgac tttctcgccg atcaaaaggc
     2281 attttgctat taagggattg acgagggcgt atctgcgcag taagatgcgc cccgcatgag
     2341 acggcatgcc gtctctagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa
     2401 cttgaaaaag tggcaccgag tcggtgcttt ttttaattcg aaaagcctgc tcaacgagca
     2461 ggcttttttg gtcgacagtt cataggtgat tgcggatccc gtcgttgaca attaatcatc
     2521 ggcatagtat atcggcatag tataatacga caaggtgagg aactaaacca tggccaagtt
     2581 gaccagtgcc gttccggtgc tcaccgcgcg cgacgtcgcc ggagcggtcg agttctggac
     2641 cgaccggctc gggttctccc gcgacttcgt ggaggacgac ttcgccggtg tggtccggga
     2701 cgacgtgacc ctgttcatca gcgcggtcca ggaccaggtg gtgccggaca acaccctggc
     2761 ctgggtgtgg gtgcgcggcc tggacgagct gtacgccgag tggtcggagg tcgtgtccac
     2821 gaacttccgg gacgcctccg ggccggccat gaccgagatc ggcgagcagc cgtgggggcg
     2881 ggagttcgcc ctgcgcgacc cggccggcaa ctgcgtgcac ttcgtggccg aggagcagga
     2941 ctgagagctc gcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg
     3001 atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat ccaagcacta
     3061 gctagtaaca acttatatcg tatggggctg acttcaggtg ctacatttga agagataaat
//
