LOCUS       Exported                6596 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPtPBR2, complete sequence.
ACCESSION   KX523202
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6596)
  AUTHORS   Diner RE, Bielinski VA, Dupont CL, Allen AE, Weyman PD.
  TITLE     Refinement of the Diatom Episome Maintenance Sequence and 
            Improvement of Conjugation-Based DNA Delivery Methods
  JOURNAL   Front Bioeng Biotechnol 4, 65 (2016)
  PUBMED    27551676
REFERENCE   2  (bases 1 to 6596)
  AUTHORS   Diner RE, Weyman PD.
  TITLE     Direct Submission
  JOURNAL   Submitted (07-JUL-2016) Scripps Institution of Oceanography, 
            University of California San Diego, 9500 Gilman Drive, La Jolla, CA 
            92093, USA
REFERENCE   3  (bases 1 to 6596)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6596
                     /organism="Cloning vector pPtPBR2"
                     /mol_type="other DNA"
                     /db_xref="taxon:1897462"
     promoter        10..38
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     regulatory      10..15
                     /regulatory_class="minus_35_signal"
                     /note="tet (P2) promoter"
     regulatory      33..38
                     /regulatory_class="minus_10_signal"
                     /note="tet (P2) promoter"
     regulatory      complement(44..49)
                     /regulatory_class="minus_10_signal"
                     /note="P1 promoter"
     regulatory      complement(64..75)
                     /regulatory_class="minus_35_signal"
                     /note="P1 promoter -35 sequence region"
     CDS             86..1276
                     /codon_start=1
                     /product="tetracycline efflux protein class C"
                     /function="confers resistance to tetracyclin"
                     /label=tetracycline efflux protein class C
                     /protein_id="AOI27796.1"
                     /translation="MKSNNALIVILGTVTLDAVGIGLVMPVLPGLLRDIVHSDSIASHY
                     GVLLALYALMQFLCAPVLGALSDRFGRRPVLLASLLGATIDYAIMATTPVLWILYAGRI
                     VAGITGATGAVAGAYIADITDGEDRARHFGLMSACFGVGMVAGPVAGGLLGAISLHAPF
                     LAAAVLNGLNLLLGCFLMQESHKGERRPMPLRAFNPVSSFRWARGMTIVAALMTVFFIM
                     QLVGQVPAALWVIFGEDRFRWSATMIGLSLAVFGILHALAQAFVTGPATKRFGEKQAII
                     AGMAADALGYVLLAFATRGWMAFPIMILLASGGIGMPALQAMLSRQVDDDHQGQLQGSL
                     AALTSLTSIIGPLIVTAIYAASASTWNGLAWIVGAALYLVCLPALRRGAWSRATST"
     regulatory      1905..1909
                     /regulatory_class="ribosome_binding_site"
                     /note="rop RBS"
     CDS             1915..2106
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(2337..2708)
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     oriT            2741..2850
                     /note="incP origin of transfer"
     primer_bind     3127..3143
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        3167..3185
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             3889..4263
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      complement(4769..5357)
                     /direction=LEFT
                     /note="/note=pMB1 origin of replication (RNAII -35 to 
                     RNA/DNA switch point)"
     misc_RNA        complement(4769..5321)
                     /label=RNAII transcript
                     /note="RNAII transcript"
     regulatory      5175..5180
                     /regulatory_class="minus_35_signal"
                     /note="RNAI promoter; TTGAAG"
     regulatory      5197..5202
                     /regulatory_class="minus_10_signal"
                     /note="RNAI promoter; GCTACA"
     misc_RNA        5211..5318
                     /label=RNAI transcript
                     /note="RNAI transcript"
     regulatory      complement(5331..5336)
                     /regulatory_class="minus_10_signal"
                     /note="RNAII promoter; CGTAAT"
     regulatory      complement(5352..5357)
                     /regulatory_class="minus_35_signal"
                     /note="RNAII promoter; TTGAGA"
     CDS             complement(5528..6388)
                     /codon_start=1
                     /gene="Bla"
                     /product="beta-lactamase"
                     /function="confers resistance to ampicillin"
                     /label=Bla
                     /protein_id="AOI27795.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(5528..6388)
                     /gene="Bla"
                     /label=Bla
     sig_peptide     complement(6320..6388)
                     /gene="Bla"
                     /note="required for secretion to the periplasm; cleaved off
                     to form the mature beta-lactamase protein"
     promoter        complement(6389..6493)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      complement(6396..6400)
                     /regulatory_class="ribosome_binding_site"
                     /note="bla RBS"
     regulatory      complement(6432..6437)
                     /regulatory_class="minus_10_signal"
                     /note="note=bla (P3) promoter; GAGACA"
     regulatory      complement(6453..6458)
                     /regulatory_class="minus_35_signal"
                     /note="bla (P3) promoter; TTCAAA"