LOCUS       Exported                8116 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pMQ150, complete sequence.
ACCESSION   EU546823
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8116)
  AUTHORS   Shanks RM, Kadouri DE, MacEachran DP, OToole GA.
  TITLE     New yeast recombineering tools for bacteria
  JOURNAL   Plasmid 62 (2), 88-97 (2009)
  PUBMED    19477196
REFERENCE   2  (bases 1 to 8116)
  AUTHORS   Shanks RM, MacEachran DP, Powers SZ, Kadouri DE, OToole GA.
  TITLE     In vivo recombination vectors for modification and expression of 
            genes in Gram-negative and Gram-positive bacteria
  JOURNAL   Unpublished
REFERENCE   3  (bases 1 to 8116)
  AUTHORS   Shanks RM, MacEachran DP, Powers SZ, Kadouri DE, OToole GA.
  TITLE     Direct Submission
  JOURNAL   Submitted (04-MAR-2008) Ophthalmology, University of Pittsburgh, 203
            Lothrop St., Pittsburgh, PA 15213, USA
REFERENCE   4  (bases 1 to 8116)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8116
                     /organism="Cloning vector pMQ150"
                     /mol_type="other DNA"
                     /label=shuttle vector
                     /note="shuttle vector"
                     /db_xref="taxon:515316"
     CDS             complement(235..1656)
                     /codon_start=1
                     /gene="sacB"
                     /product="SacB"
                     /label=sacB
                     /note="sucrose sensitivity marker"
                     /protein_id="ACB45216.1"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     gene            complement(235..1656)
                     /gene="sacB"
                     /label=sacB
     promoter        complement(1657..2102)
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     oriT            2130..2954
                     /note="oriT"
     oriT            2476..2585
                     /note="incP origin of transfer"
     regulatory      complement(3006..3208)
                     /regulatory_class="terminator"
     terminator      3006..3033
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      3125..3211
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(3367..3651)
                     /codon_start=1
                     /gene="lacZalpha"
                     /product="beta-gal alpha peptide"
                     /label=lacZalpha
                     /protein_id="ACB45217.1"
                     /translation="MTMITNSSSVPGDPLESTCRHASLALAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRAWLFWRMREDFQPDTD"
     gene            complement(3367..3651)
                     /gene="lacZalpha"
                     /label=lacZalpha
     primer_bind     3561..3577
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3581..3637
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(3647..3663)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    complement(3669..3689)
                     /label=lacO
                     /note="lacO"
     protein_bind    3671..3687
                     /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(3695..3725)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3740..3761
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     regulatory      complement(3773..3960)
                     /regulatory_class="promoter"
                     /note="Plac"
     promoter        complement(3812..3830)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      4038..4426
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     rep_origin      4098..4318
                     /label=oriR6K-gamma
                     /note="oriR6K-gamma"
     CDS             4777..5571
                     /codon_start=1
                     /gene="nptII"
                     /product="aminoglycoside 3-phosphotransferase"
                     /label=nptII
                     /note="confers resistance to kanamycin; KanR"
                     /protein_id="ACB45218.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            4777..5571
                     /gene="nptII"
                     /label=nptII
     CDS             5868..6242
                     /codon_start=1
                     /gene="rpsL"
                     /product="ribosomal protein L"
                     /label=rpsL
                     /note="strep sensitivity counter-selectable marker"
                     /protein_id="ACB45219.1"
                     /translation="MATVNQLVRKPRARKVAKSNVPALEACPQKRGVCTRVYTTTPKKP
                     NSALRKVCRVRLTNGFEVTSYIGGEGHNLQEHSVILIRGGRVKDLPGVRYHTVRGALDC
                     SGVKDRKQARSKYGVKRPKA"
     gene            5868..6242
                     /gene="rpsL"
                     /label=rpsL
     CDS             complement(6502..7305)
                     /codon_start=1
                     /gene="URA3"
                     /product="Ura3"
                     /label=URA3
                     /note="uracil biosynthesis protein"
                     /protein_id="ACB45220.1"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     gene            complement(6502..7305)
                     /gene="URA3"
                     /label=URA3
     promoter        complement(7306..7521)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     rep_origin      7563..8076
                     /label=CEN6/ARSH4 derived from Saccharomyces cerevisiae
                     /note="CEN6/ARSH4 derived from Saccharomyces cerevisiae"
     misc_feature    7573..8076
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"