LOCUS       Exported                6806 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Shuttle vector pP01680-nat-cel8A, complete sequence.
ACCESSION   MH328018
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6806)
  AUTHORS   Heinze S, Zimmermann K, Ludwig C, Heinzlmeir S, Schwarz WH, Zverlov 
            VV, Liebl W, Kornberger P.
  TITLE     Efficient secretory heterologous enzyme production in the novel host
            Paenibacillus polymyxa by usage of its promoter sequences
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6806)
  AUTHORS   Heinze S, Zimmermann K, Ludwig C, Heinzlmeir S, Schwarz WH, Zverlov 
            VV, Liebl W, Kornberger P.
  TITLE     Direct Submission
  JOURNAL   Submitted (09-MAY-2018) Department for Microbiology, Technical 
            University of Munich, Emil-Ramann-Str. 4, Freising, Bavaria 85354, 
            Germany
REFERENCE   3  (bases 1 to 6806)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6806
                     /organism="Shuttle vector pP01680-nat-cel8A"
                     /mol_type="other DNA"
                     /label=derived from Escherichia coli TOP10
                     /note="derived from Escherichia coli TOP10"
                     /db_xref="taxon:2282828"
     regulatory      complement(77..676)
                     /regulatory_class="replication_regulatory_region"
                     /note="ColE1 origin of replication, E. coli"
     rep_origin      complement(79..667)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(838..1698)
                     /codon_start=1
                     /gene="ampR"
                     /product="AmpR"
                     /label=ampR
                     /note="Ampicillin Resistance"
                     /protein_id="AXF54519.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(838..1698)
                     /gene="ampR"
                     /label=ampR
     promoter        complement(1699..1803)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      complement(1832..2353)
                     /regulatory_class="other"
                     /note="origin of transfer for conjugation, contains traJ 
                     and oriT"
     CDS             complement(1832..2116)
                     /codon_start=1
                     /gene="traJ"
                     /product="TraJ"
                     /label=traJ
                     /protein_id="AXF54520.1"
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGQQKAHYVNHHPNQVFWGRGAVKH"
     gene            complement(1832..2116)
                     /gene="traJ"
                     /label=traJ
     regulatory      complement(2117..2353)
                     /regulatory_class="other"
                     /note="oriT (origin of transfer)"
     oriT            2149..2258
                     /note="incP origin of transfer"
     CDS             complement(2515..3276)
                     /codon_start=1
                     /gene="kanR"
                     /product="KanR"
                     /label=kanR
                     /note="Kanamycin Resistance"
                     /protein_id="AXF54521.1"
                     /translation="MNGPIIMTREERMKIVHEIKERILDKYGDDVKAIGVYGSLGRQTD
                     GPYSDIEMMCVMSTEEAEFSHEWTTGEWKVEVNFDSEEILLDYASQVESDWPLTHGQFF
                     SILPIYDSGGYLEKVYQTAKSVEAQTFHDAICALIVEELFEYAGKWRNIRVQGPTTFLP
                     SLTVQVAMAGAMLIGLHHRICYTTSASVLTEAVKQSDLPSGYDHLCQFVMSGQLSDSEK
                     LLESLENFWNGIQEWTERHGYIVDVSKRIPF"
     gene            complement(2515..3276)
                     /gene="kanR"
                     /label=kanR
     regulatory      complement(3445..4612)
                     /regulatory_class="replication_regulatory_region"
                     /note="pUB origin of replication, Bacillus"
     CDS             complement(3445..4449)
                     /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"
     regulatory      4866..5158
                     /regulatory_class="promoter"
                     /note="P01680 promoter from P. polymyxa DSM292"
     gene            5156..6646
                     /gene="SPnat-Cel8A-His6"
                     /label=SPnat-Cel8A-His6
     CDS             5156..5267
                     /codon_start=1
                     /gene="SPnat-Cel8A-His6"
                     /product="signal peptide of P. polymyxa DSM292 ORF 
                     PPOLYM_01680"
                     /label=SPnat-Cel8A-His6
                     /protein_id="AXF54522.1"
                     /translation="MKSFLVKSLLALSVLFMLLGFMPVGQASVSANSVGAE"
     CDS             5285..6646
                     /codon_start=1
                     /gene="SPnat-Cel8A-His6"
                     /product="Cel8A with C-terminal hexahistidine-tag"
                     /label=SPnat-Cel8A-His6
                     /protein_id="AXF54523.1"
                     /translation="AGVPFNTKYPYGPTSIADNQSEVTAMLKAEWEDWKSKRITSNGAG
                     GYKRVQRDASTNYDTVSEGMGYGLLLAVCFNEQALFDDLYRYVKSHFNGNGLMHWHIDA
                     NNNVTSHDGGDGAATDADEDIALALIFADKLWGSSGAINYGQEARTLINNLYNHCVEHG
                     SYVLKPGDRWGGSSVTNPSYFAPAWYKVYAQYTGDTRWNQVADKCYQIVEEVKKYNNGT
                     GLVPDWCTASGTPASGQSYDYKYDATRYGWRTAVDYSWFGDQRAKANCDMLTKFFARDG
                     AKGIVDGYTIQGSKISNNHNASFIGPVAAASMTGYDLNFAKELYRETVAVKDSEYYGYY
                     GNSLRLLTLLYITGNFPNPLSDLSGQPTPPSNPTPSLPPQVVYGDVNGDGNVNSTDLTM
                     LKRYLLKSVTNINREAADVNRDGAINSSDMTILKRYLIKSIPHLPYSRHHHHHH"
     CDS             6626..6643
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"