LOCUS       Exported                4961 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Transposon donor vector pRZ49, complete sequence.
ACCESSION   KP698385
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4961)
  AUTHORS   Zordan RE, Beliveau BJ, Trow JA, Craig NL, Cormack BP.
  TITLE     Avoiding the ends: internal epitope tagging of proteins using 
            transposon Tn7
  JOURNAL   Genetics 200 (1), 47-58 (2015)
  PUBMED    25745023
REFERENCE   2  (bases 1 to 4961)
  AUTHORS   Zordan RE, Cormack BP.
  TITLE     Direct Submission
  JOURNAL   Submitted (24-JAN-2015) Molecular Biology and Genetics, Johns 
            Hopkins School of Medicine, 725 N Wolfe St., Hunterian 609, 
            Baltimore, MD 21205, USA
REFERENCE   3  (bases 1 to 4961)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..4961
                     /organism="Transposon donor vector pRZ49"
                     /mol_type="other DNA"
                     /db_xref="taxon:1631654"
     CDS             361..1155
                     /codon_start=1
                     /gene="kanR"
                     /product="aminoglycoside-3-O-phosphotransferase"
                     /label=kanR
                     /note="kanamycin resistance protein"
                     /protein_id="AKA28276.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            361..1155
                     /gene="kanR"
                     /label=kanR
     protein_bind    1430..1451
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        1466..1496
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1504..1520
                     /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)."
     primer_bind     1528..1544
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     mobile_element  1597..3442
                     /mobile_element_type="transposon:Tn7-mEOS2"
     misc_feature    1820..1849
                     /label=linker 1
                     /note="linker 1"
     misc_feature    1856..2533
                     /label=similar to mEOS2 fluorescent protein
                     /note="similar to mEOS2 fluorescent protein"
     CDS             1856..2533
                     /codon_start=1
                     /product="monomeric variant of green-to-red photoswitchable
                     fluorescent protein EosFP (McKinney et al., 2009)"
                     /label=mEos2
                     /translation="MSAIKPDMKIKLRMEGNVNGHHFVIDGDGTGKPFEGKQSMDLEVK
                     EGGPLPFAFDILTTAFHYGNRVFAKYPDNIQDYFKQSFPKGYSWERSLTFEDGGICIAR
                     NDITMEGDTFYNKVRFYGTNFPANGPVMQKKTLKWEPSTEKMYVRDGVLTGDIHMALLL
                     EGNAHYRCDFRTTYKAKEKGVKLPGYHFVDHCIEILSHDKDYNKVKLYEHAVAHSGLPD
                     NARR"
     misc_feature    2534..2569
                     /label=linker 2
                     /note="linker 2"
     misc_feature    2585..3365
                     /label=dhfr gene cassette
                     /note="dhfr gene cassette"
     CDS             2763..3221
                     /codon_start=1
                     /gene="dhfr"
                     /product="dihydrofolate reductase"
                     /label=dhfr
                     /protein_id="AKA28277.1"
                     /translation="MVAISKNGVIGNGPDIPWSAKGEQLLFKAITYNQWLLVGRKTFGS
                     MGALPNRKYAVVTRSSFTSDNENVLIFPSIKDALTNLKKITDHVIVSGGGEIYKSLIDQ
                     VDTLHISTIDIEPEGDVYFPEIPSNFRPVFTQDFASNINYSYQIWQKG"
     gene            2763..3221
                     /gene="dhfr"
                     /label=dhfr
     primer_bind     complement(3449..3465)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3771..4544
                     /label=mob RP4
                     /note="mob RP4"
     CDS             complement(3897..4268)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            4301..4410
                     /note="incP origin of transfer"
     rep_origin      4570..4961
                     /label=R6K gamma
                     /note="R6K gamma"
     rep_origin      4570..4958
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"