LOCUS       Exported                6668 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Plasmid vector pSS4332, complete sequence.
ACCESSION   KT823413
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6668)
  AUTHORS   Plaut RD, Stibitz S.
  TITLE     Improvements to a Markerless Allelic Exchange System for Bacillus 
            anthracis
  JOURNAL   PLoS ONE 10 (12), E0142758 (2015)
  PUBMED    26624016
REFERENCE   2  (bases 1 to 6668)
  AUTHORS   Plaut RD, Stibitz S.
  TITLE     Direct Submission
  JOURNAL   Submitted (24-SEP-2015) Center for Biologics Evaluation and 
            Research, OVRR, DBPAP, Food and Drug Administration, 10903 New 
            Hampshire Ave, Bldg 52/72, Rm 3310, Silver Spring, MD 20993, USA
REFERENCE   3  (bases 1 to 6668)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6668
                     /organism="Plasmid vector pSS4332"
                     /mol_type="other DNA"
                     /db_xref="taxon:1764030"
     CDS             complement(13..714)
                     /codon_start=1
                     /gene="I-SceI"
                     /product="I-SceI"
                     /label=I-SceI
                     /protein_id="ALQ43917.1"
                     /translation="MHQKNQVMNLGPNSKLLKEYKSQLIELNIEQFEAGIGLILGDAYI
                     RSRDEGKTYCMQFEWKNKAYMDHVCLLYDQWVLSPPHKKERVNHLGNLVITWGAQTFKH
                     QAFNKLANLFIVNNKKTIPNNLVENYLTPMSLAYWFMDDGGKWDYNKNSTNKSIVLNTQ
                     SFTFEEVEYLVKGLRNKFQLNCYVKINKNKPIIYIDSMSYLIFYNLIKPYLIPQMMYKL
                     PNTISSETFLK"
     gene            complement(13..714)
                     /gene="I-SceI"
                     /label=I-SceI
     CDS             complement(764..1453)
                     /codon_start=1
                     /gene="AmCyan"
                     /product="AmCyan"
                     /label=AmCyan
                     /protein_id="ALQ43918.1"
                     /translation="MALSNKFIGDDMKMTYHMDGCVNGHYFTVKGEGSGKPYEGTQTST
                     FKVTMANGGPLAFSFDILSTVFMYGNRCFTAYPTSMPDYFKQAFPDGMSYERTFTYEDG
                     GVATASWEISLKGNCFEHKSTFHGVNFPADGPVMAKMTTGWDPSFEKMTVCDGILKGDV
                     TAFLMLQGGGNYRCQFHTSYKTKKPVTMPPNHAVEHRIARTDLDKGGNSVQLTEHAVAH
                     ITSVVPF"
     gene            complement(764..1453)
                     /gene="AmCyan"
                     /label=AmCyan
     regulatory      complement(1480..1485)
                     /regulatory_class="minus_10_signal"
                     /note="PFP1"
     regulatory      complement(1503..1508)
                     /regulatory_class="minus_35_signal"
                     /note="PFP1"
     CDS             complement(1626..2315)
                     /codon_start=1
                     /gene="AmCyan"
                     /product="AmCyan"
                     /label=AmCyan
                     /note="tandem repeat"
                     /protein_id="ALQ43919.1"
                     /translation="MALSNKFIGDDMKMTYHMDGCVNGHYFTVKGEGSGKPYEGTQTST
                     FKVTMANGGPLAFSFDILSTVFMYGNRCFTAYPTSMPDYFKQAFPDGMSYERTFTYEDG
                     GVATASWEISLKGNCFEHKSTFHGVNFPADGPVMAKMTTGWDPSFEKMTVCDGILKGDV
                     TAFLMLQGGGNYRCQFHTSYKTKKPVTMPPNHAVEHRIARTDLDKGGNSVQLTEHAVAH
                     ITSVVPF"
     gene            complement(1626..2315)
                     /gene="AmCyan"
                     /label=AmCyan
                     /note="tandem repeat"
     regulatory      complement(2342..2347)
                     /regulatory_class="minus_10_signal"
                     /note="PFP1"
     regulatory      complement(2365..2370)
                     /regulatory_class="minus_35_signal"
                     /note="PFP1"
     CDS             complement(2377..3171)
                     /codon_start=1
                     /gene="kan"
                     /gene_synonym="aphA(3)"
                     /product="kan"
                     /label=kan
                     /note="from Staphylococcus aureus"
                     /protein_id="ALQ43920.1"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     gene            complement(2377..3171)
                     /gene="kan"
                     /gene_synonym="aphA(3)"
                     /label=kan
                     /note="from Staphylococcus aureus"
     promoter        complement(3473..3491)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(3512..3528)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3536..3552
                     /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(3560..3590)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3605..3626
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      3858..4525
                     /label=pUC ori
                     /note="pUC ori"
     rep_origin      complement(3914..4502)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     oriT            4697..4948
                     /label=from D2BpUC9Cam
                     /note="from D2BpUC9Cam"
     oriT            4840..4949
                     /note="incP origin of transfer"
     oriT            4955..5206
                     /note="from D2BpUC9Cam; tamdem repeat"
     oriT            5098..5207
                     /note="incP origin of transfer"
     CDS             complement(5404..6408)
                     /codon_start=1
                     /gene="repB"
                     /product="RepB replication protein"
                     /label=repB
                     /note="from Enterococcus faecalis plasmid pAM-alpha-1"
                     /db_xref="GI:22652809"
                     /protein_id="AAN03827.1"
                     /translation="MGVSFNIMCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDY
                     FELLHILEFKKAERVKDCAEILEYKQNRETGERKLYRVWFCKSRLCPMCNWRRAMKHGI
                     QSQKVVAEVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLV
                     GFMRATEVTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPN
                     VKVQMIRPKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLIS
                     YGGLLKEIHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
     CDS             complement(5404..6387)
                     /codon_start=1
                     /gene="repU"
                     /product="RepU"
                     /label=repU
                     /note="from pBC16"
                     /protein_id="ALQ43921.1"
                     /translation="MCPNSSIYSDEKSRVLVDKTKSGKVRPWREKKIANVDYFELLHIL
                     EFKKAERVKDCAEILEYKQNRETGERKLYRVWFCKSRLCPMCNWRRAMKHGIQSQKVVA
                     EVIKQKPTVRWLFLTLTVKNVYDGEELNKSLSDMAQGFRRMMQYKKINKNLVGFMRATE
                     VTINNKDNSYNQHMHVLVCVEPTYFKNTENYVNQKQWIQFWKKAMKLDYDPNVKVQMIR
                     PKNKYKSDIQSAIDETAKYPVKDTDFMTDDEEKNLKRLSDLEEGLHRKRLISYGGLLKE
                     IHKKLNLDDTEEGDLIHTDDDEKADEDGFSIIAMWNWERKNYFIKE"
     gene            complement(5404..6387)
                     /gene="repU"
                     /label=repU
                     /note="from pBC16"