LOCUS       Exported                7163 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pR6K-lys-Gm, complete sequence.
ACCESSION   EF153730
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7163)
  AUTHORS   Kang Y, Son MS, Hoang TT.
  TITLE     One step engineering of T7-expression strains for protein 
            production: increasing the host-range of the T7-expression system
  JOURNAL   Protein Expr. Purif. 55 (2), 325-333 (2007)
  PUBMED    17716915
REFERENCE   2  (bases 1 to 7163)
  AUTHORS   Kang Y, Son MS, Hoang TT.
  TITLE     Direct Submission
  JOURNAL   Submitted (30-NOV-2006) Microbiology, University of Hawaii at Manoa,
            2538 The Mall - Snyder 310, Honolulu, HI 96822, USA
REFERENCE   3  (bases 1 to 7163)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7163
                     /organism="Expression vector pR6K-lys-Gm"
                     /mol_type="other DNA"
                     /db_xref="taxon:418829"
     misc_feature    7..25
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
     protein_bind    37..268
                     /gene="attP"
                     /label=-lambda attP
                     /bound_moiety="lambda integrase"
                     /note="integrase from phage lambda"
     rep_origin      587..945
                     /label=R6K-gamma-ori
                     /note="R6K-gamma-ori"
     misc_feature    951..969
                     /label=Tn5 ME
                     /note="hyperactive mosaic end for Tn5 transposase 
                     recognition (Reznikoff et al., 2004)"
     CDS             complement(978..3629)
                     /codon_start=1
                     /gene="T7-pol"
                     /product="T7 RNA polymerase"
                     /label=T7-pol
                     /protein_id="ABN54763.1"
                     /translation="MNTINIAKNDFSDIELAAIPFNTLADHYGERLAREQLALEHESYE
                     MGEARFRKMFERQLKAGEVADNAAAKPLITTLLPKMIARINDWFEEVKAKRGKRPTAFQ
                     FLQEIKPEAVAYITIKTTLACLTSADNTTVQAVASAIGRAIEDEARFGRIRDLEAKHFK
                     KNVEEQLNKRVGHVYKKAFMQVVEADMLSKGLLGGEAWSSWHKEDSIHVGVRCIEMLIE
                     STGMVSLHRQNAGVVGQDSETIELAPEYAEAIATRAGALAGISPMFQPCVVPPKPWTGI
                     TGGGYWANGRRPLALVRTHSKKALMRYEDVYMPEVYKAINIAQNTAWKINKKVLAVANV
                     ITKWKHCPVEDIPAIEREELPMKPEDIDMNPEALTAWKRAAAAVYRKDKARKSRRISLE
                     FMLEQANKFANHKAIWFPYNMDWRGRVYAVSMFNPQGNDMTKGLLTLAKGKPIGKEGYY
                     WLKIHGANCAGVDKVPFPERIKFIEENHENIMACAKSPLENTWWAEQDSPFCFLAFCFE
                     YAGVQHHGLSYNCSLPLAFDGSCSGIQHFSAMLRDEVGGRAVNLLPSETVQDIYGIVAK
                     KVNEILQADAINGTDNEVVTVTDENTGEISEKVKLGTKALAGQWLAYGVTRSVTKRSVM
                     TLAYGSKEFGFRQQVLEDTIQPAIDSGKGLMFTQPNQAAGYMAKLIWESVSVTVVAAVE
                     AMNWLKSAAKLLAAEVKDKKTGEILRKRCAVHWVTPDGFPVWQEYKKPIQTRLNLMFLG
                     QFRLQPTINTNKDSEIDAHKQESGIAPNFVHSQDGSHLRKTVVWAHEKYGIESFALIHD
                     SFGTIPADAANLFKAVRETMVDTYESCDVLADFYDQFADQLHESQLDKMPALPAKGNLN
                     LRDILESDFAFA"
     gene            complement(978..3629)
                     /gene="T7-pol"
                     /label=T7-pol
     CDS             complement(3626..4111)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGEWRFAWFPAPEAVPESWLECDLPEADTVVVPSNWQMHGYDAPIYTNV
                     TYPITVNPPFVPTENPTGCYSLTFNVDESWLQEGQTRIIFDGVRDLIPDLLTGRGTK"
     primer_bind     4075..4091
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4107..4123)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    complement(4129..4153)
                     /label=lacUV5 operator
                     /note="lacUV5 operator"
     protein_bind    4131..4147
                     /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(4155..4185)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     regulatory      complement(4156..4185)
                     /regulatory_class="promoter"
                     /note="lacUV5 promoter"
     protein_bind    4200..4221
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(4234..5316)
                     /codon_start=1
                     /gene="lacIq"
                     /product="LacI"
                     /label=lacIq
                     /note="lac repressor"
                     /protein_id="ABN54764.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPSTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     gene            complement(4234..5316)
                     /gene="lacIq"
                     /label=lacIq
     misc_feature    complement(5397..5444)
                     /label=Flp-recombination-target
                     /note="Flp-recombination-target"
     protein_bind    complement(5397..5444)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        5485..5513
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             5702..6235
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyl transferase"
                     /function="gentamycin resistance"
                     /label=aacC1
                     /protein_id="ABN54765.1"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     gene            5702..6235
                     /gene="aacC1"
                     /label=aacC1
     misc_feature    complement(6365..6412)
                     /label=Flp-recombination-target
                     /note="Flp-recombination-target"
     protein_bind    complement(6365..6412)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             6675..7130
                     /codon_start=1
                     /gene="T7-lys"
                     /product="T7 lysozyme"
                     /label=T7-lys
                     /note="native inhibitor of T7 RNA polymerase"
                     /protein_id="ABN54766.1"
                     /translation="MARVQFKQRESTDAIFVHCSATKPSQNVGVREIRQWHKEQGWLDV
                     GYHFIIKRDGTVEAGRDEMAVGSHAKGYNHNSIGVCLVGGIDDKGKFDANFTPAQMQSL
                     RSLLVTLLAKYEGAGLRAHHEVAPKACPSFDLKRWWEKNELVTSDRG"
     gene            6675..7130
                     /gene="T7-lys"
                     /label=T7-lys