LOCUS       Exported                5004 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pARKANI, complete sequence.
ACCESSION   MG100204
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5004)
  AUTHORS   Eguia FAP., Ramos HR, Kraschowetz S, Omote DQ, Ramos CRR., Ho PL, 
            Carvalho E, Goncalves VM.
  TITLE     A new vector for heterologous gene expression in Escherichia coli 
            with increased stability in the absence of antibiotic
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5004)
  AUTHORS   Eguia FAP., Ramos HR, Kraschowetz S, Omote DQ, Ramos CRR., Ho PL, 
            Carvalho E, Goncalves VM.
  TITLE     Direct Submission
  JOURNAL   Submitted (06-OCT-2017) Centro de Biotecnologia, Instituto Butantan,
            Av Vital Brasil, 1500, Sao Paulo, SP 05503-900, Brazil
REFERENCE   3  (bases 1 to 5004)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Assembly Method       :: CLC Genomics Workbench v. 9.0
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5004
                     /organism="Expression vector pARKANI"
                     /mol_type="other DNA"
                     /db_xref="taxon:2116677"
     RBS             11..33
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    48..72
                     /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        complement(73..91)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        400..477
                     /gene="lacI"
                     /label=lacI promoter
     CDS             601..969
                     /codon_start=1
                     /gene="lacI"
                     /product="Lac repressor"
                     /label=lacI
                     /protein_id="AVN68413.1"
                     /translation="MAELNYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSR
                     ADQLGASVVVSMVERSGVEACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPA
                     LFLDVSDQTPINSIIFLP"
     gene            601..969
                     /gene="lacI"
                     /label=lacI
     misc_feature    complement(1946..2314)
                     /label=plasmid partitioning sequence - par
                     /note="plasmid partitioning sequence - par"
     primer_bind     complement(2356..2372)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2380..2396
                     /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        complement(2404..2434)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2449..2470
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(2759..3349)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(3634..4428)
                     /codon_start=1
                     /gene="kmr"
                     /product="kanamycin resistance protein"
                     /label=kmr
                     /note="APH"
                     /protein_id="AVN68412.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            complement(3634..4428)
                     /gene="kmr"
                     /label=kmr
     terminator      4823..4870
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(4987..5004)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"