pGEX_LacI_TetR_chPylRS_IPYE vector (V016348)

LOCUS       62056_11515        5990 bp DNA     circular SYN 19-APR-2020
DEFINITION  Cloning vector pGEX_LacI_TetR_chPylRS_IPYE, complete sequence.
ACCESSION   MN882189
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5990)
  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 5990)
  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 5990)
  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..5990
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      19..607
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     regulatory      717..786
                     /label=AmpR promoter
                     /note="AmpR promoter"
                     /regulatory_class="promoter"
     CDS             846..1466
                     /label=TetR
                     /note="tetracycline repressor TetR"
     terminator      complement(1491..1577)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     promoter        1893..1970
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             1971..3050
                     /label=lacI
                     /note="lac repressor"
     promoter        3171..3199
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     protein_bind    3207..3223
                     /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)."
     CDS             3246..4505
                     /codon_start=1
                     /transl_table=11
                     /product="chimeric PylRS variant"
                     /label=chimeric PylRS variant
                     /note="chimeric PylRS variant; Methanosarcina mazei/barkeri
                     chPylRS(IPYE)"
                     /protein_id="QIZ64217.1"
                     /translation="MDKKPLDVLISATGLWMSRTGTLHKIKHYEISRSKIYIEMACGDH
                     LVVNNSRSCRPARAFRYHKYRKTCKRCRVSDEDINNFLTRSTEGKTSVKVKVVSEPKVK
                     KAMPKSVSRAPKPLENPVSAKASTDTSRSVPSPAKSTPNSPVPTSASAPALTKSQTDRL
                     EVLLNPKDEISLNSGKPFRELESELLSRRKKDLQQIYAEERENYLGKLEREITRFFVDR
                     GFLEIKSPILIPLEYIERMGIDNDTELSKQIFRVDKNFCLRPMLAPNLYNYLRKLDRAL
                     PDPIKIFEIGPCYRKESDGKEHLEEFTMLNFCQMGSGCTRENLESIITDFLNHLGIDFK
                     IVGDSCMVYGDTLDVMHGDLELSSAVVGPIPLDREWGIDKPWIGAGFGLERLLKVKHDF
                     KNIKRAARSESYYNGISTNL"
     terminator      4729..4776
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        4873..4977
                     /label=AmpR promoter
     CDS             4978..5835
                     /label=AmpR
                     /note="beta-lactamase"