LOCUS       Exported                6207 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Broad host range vector vector pCVD076, complete sequence.
ACCESSION   KM017876
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6207)
  AUTHORS   Taton A, Unglaub F, Wright NE, Zeng WY, Paz-Yepez J, Brahamsha B, 
            Palenik B, Peterson TC, Haerizadeh F, Golden SS, Golden JW.
  TITLE     Broad-host-range vector system for synthetic biology and 
            biotechnology in cyanobacteria
  JOURNAL   Nucleic Acids Res. (2014) In press
  PUBMED    25074377
REFERENCE   2  (bases 1 to 6207)
  AUTHORS   Taton A, Unglaub F, Wright N, Zeng WY, Paz Yepes J, Brahamsha B, 
            Palenik B, Peterson T, Haerizadeh F, Golden SS, Golden JW.
  TITLE     Direct Submission
  JOURNAL   Submitted (17-JUN-2014) Division of Biological Sciences, University 
            of California, San Diego, 9500 Gilman Dr., La Jolla, CA 92093, USA
REFERENCE   3  (bases 1 to 6207)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6207
                     /organism="Broad host range vector vector pCVD076"
                     /mol_type="other DNA"
                     /note="A7120NS2 is a chromosome integration device that 
                     comprises the recombination site NS2 for Anabaena sp 
                     PCC7120, an origin of replication for E coli, a bom site, 
                     for conjugal transfer, from pBR322 and the sacB gene for 
                     selection of double recombinants grown with sucrose A point
                     mutation was identified (G249C) in the ORF of sacB leading 
                     to the following amino acid change: E83D (Ryan Simkovsky) 
                     This device appears however functional in Anabaena sp 
                     PCC7120 and Synechococcus elongatus PCC7942"
                     /db_xref="taxon:1531930"
     misc_feature    1..6
                     /label=ZraI
                     /note="ZraI"
     misc_feature    4..4978
                     /note="CVD-BHR-NS; device for the GC-adaptor assembly: BHR 
                     replicon and neutral sites or right arm of a double 
                     recombination KO"
     misc_feature    4..24
                     /note="G5C5; GC-adaptor for seamless assembly, flanking a 
                     BHR replicons, neutral sites, or cyanobacterial replicons; 
                     synthetic"
     misc_feature    25..30
                     /label=XbaI
                     /note="XbaI"
     misc_recomb     complement(31..1035)
                     /note="A7120NS2-LA; left arm of the homologous 
                     recombination site NS2 (all1997_D43-all1697_D1) into the 
                     chromosome of Anabaena sp. PCC7120 derived from INSD 
                     accession NC_003272"
     rep_origin      1046..1719
                     /note="ori-pBR322; origin of replication of pBR322"
     rep_origin      1072..1660
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    1846..1986
                     /label=bom
                     /note="basis of mobility region from pBR322"
     oriT            1892..2005
                     /note="bom; bom site of pBR322, for comjugal transfer; 
                     requires a helper plasmid such as pRL623 (or pRK2013)"
     misc_feature    2149..2154
                     /label=EcoRV
                     /note="EcoRV"
     CDS             complement(2174..3595)
                     /codon_start=1
                     /product="sacB"
                     /label=sacB
                     /note="sacB derived from Bacillus subtilis INSD accession 
                     AAA72307; 1; a point mutation was identified G249C in the 
                     ORF of sacB leading to the following amino acid change: 
                     E83D Ryan Simkovsky; this device appears however functional
                     in Anabaena sp. PCC7120 and Synechococcus elongatus PCC7942
                     Bacillus subtilis"
                     /protein_id="AII99642.1"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYQVPEFDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     regulatory      complement(3596..3901)
                     /regulatory_class="promoter"
                     /note="PsacB; derived from Bacillus subtilis INSD accession
                     AAA72307"
     misc_recomb     complement(3902..4951)
                     /note="A7120NS2-RA; right arm of the homologous 
                     recombination site NS2 (all1997_D43-all1697_D1) into the 
                     chromosome of Anabaena sp. PCC7120 derived from INSD 
                     accession NC_003272"
     misc_feature    4952..4957
                     /label=NheI
                     /note="NheI"
     misc_feature    4958..4978
                     /note="GC; GC-adaptor for seamless assembly, flanking BHR 
                     replicons or neutral sites; synthetic"
     misc_feature    4976..4981
                     /label=ZraI
                     /note="ZraI"
     CDS             complement(5102..5962)
                     /codon_start=1
                     /product="bla"
                     /label=bla
                     /note="confers ampicillin resistance; beta-lactamase; 
                     derived from pUC19"
                     /protein_id="AII99641.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(5963..6067)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      complement(5963..6032)
                     /regulatory_class="promoter"
                     /note="Pbla; upstream region of the bla ORF including 
                     promoter and RBS; derived from pUC19"
     misc_feature    6093..6098
                     /label=ZraI
                     /note="ZraI"