LOCUS       Exported                7140 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Broad host range vector vector pCVD068, complete sequence.
ACCESSION   KM017871
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7140)
  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 7140)
  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 7140)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7140
                     /organism="Broad host range vector vector pCVD068"
                     /mol_type="other DNA"
                     /note="RSF1010Y25FK14* is a replication device carrying a 
                     broad host range origin of replication, an origin of 
                     transfer, the rep and mob genes derived from RSF1010 mobA 
                     and mobC carries the point mutations, Y25F and K14*, 
                     respectively that prevent self-mobilization and possibly 
                     increase plasmid copy number"
                     /db_xref="taxon:1531924"
     CDS             complement(121..981)
                     /codon_start=1
                     /product="bla"
                     /label=bla
                     /note="confers ampicillin resistance; beta-lactamase; 
                     derived from pUC19"
                     /protein_id="AII99624.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(982..1086)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      complement(982..1051)
                     /regulatory_class="promoter"
                     /note="Pbla; upstream region of the bla ORF including 
                     promoter and RBS; derived from pUC19"
     misc_feature    1112..1117
                     /label=ZraI
                     /note="ZraI"
     misc_feature    1227..1232
                     /label=ZraI
                     /note="ZraI"
     misc_feature    1230..7137
                     /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    1230..1250
                     /note="G5C5; GC-adaptor for seamless assembly, flanking a 
                     BHR replicons, neutral sites, or cyanobacterial replicons; 
                     synthetic"
     misc_feature    1251..1256
                     /label=XbaI
                     /note="XbaI"
     regulatory      1288..1314
                     /regulatory_class="terminator"
                     /note="trpA T; trpA terminator from Lorist6 INSD accession 
                     X98450, one nt shorter than reference; Lorist6"
     CDS             complement(1647..2498)
                     /codon_start=1
                     /product="repC"
                     /label=repC
                     /note="repC-RSF1010; replication gene; derived from 
                     pRL1383a in INSD accession AF403426, a derivative of 
                     RSF1010 INSD accession M28829"
                     /protein_id="AII99630.1"
                     /translation="MVKPKNKHSLSHVRHDPAHCLAPGLFRALKRGERKRSKLDVTYDY
                     GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKWEAV
                     TAECHVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRKRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     CDS             complement(2485..3324)
                     /codon_start=1
                     /product="repA"
                     /label=repA
                     /note="repA-RSF1010; replication gene; derived from 
                     pRL1383a in INSD accession AF403426, a derivative of 
                     RSF1010 INSD accession M28829"
                     /protein_id="AII99629.1"
                     /translation="MATHKPINILEAFAAAPPPLDYVLPNMVAGTVGALVSPGGAGKSM
                     LALQLAAQIAGGPDLLEVGELPTGPVIYLPAEDPPTAIHHRLHALGAHLSAEERQAVAD
                     GLLIQPLIGSLPNIMAPEWFDGLKRAAEGRRLMVLDTLRRFHIEEENASGPMAQVIGRM
                     EAIAADTGCSIVFLHHASKGAAMMGAGDQQQASRGSSVLVDNIRWQSYLSSMTSAEAEE
                     WGVDDDQRRFFVRFGVSKANYGAPFADRWFRRHDGGVLKPAVLERQRKSKGVPRGEA"
     CDS             complement(3354..3560)
                     /codon_start=1
                     /product="repF"
                     /label=repF
                     /note="repF-RSF1010; replication gene; derived from 
                     pRL1383a in INSD accession AF403426, a derivative of 
                     RSF1010 INSD accession M28829"
                     /protein_id="AII99627.1"
                     /translation="MKDQKDKQTGDLLASPDAVRQARYAERMKAKGMRQRKFWLTDDEY
                     EALRECLEELRAAQGGGSDPASA"
     CDS             complement(3562..3774)
                     /codon_start=1
                     /product="repE"
                     /label=repE
                     /note="repE-RSF1010; replication gene; derived from 
                     pRL1383a in INSD accession AF403426, a derivative of 
                     RSF1010 INSD accession M28829"
                     /protein_id="AII99628.1"
                     /translation="MEYEKSASGSVYLIKSDKGYWLPGGFGYTSNKAEAGRFSVADMAS
                     LNLDGCTLSLFREDKPFGPGKFLGD"
     regulatory      complement(3804..3831)
                     /regulatory_class="promoter"
                     /note="P4; derived from pRL1383a in INSD accession 
                     AF403426, a derivative of RSF1010 INSD accession M28829; 
                     the P4 promoter is under the auto-regulated control of the 
                     F repressor, whose gene is located in the same operon, 
                     providing transcription of E-F-repA-repC-operon"
     CDS             complement(3835..5964)
                     /codon_start=1
                     /product="mobA_Y25F"
                     /label=mobA_Y25F
                     /note="mobilization gene; derived from pRL1383a in INSD 
                     accession AF403426, a derivative of RSF1010 INSD accession 
                     M28829, point mutation Y25F to reduce the plasmid mobility 
                     by more than 99%"
                     /protein_id="AII99626.1"
                     /translation="MAIYHLTAKTGSRSGGQSARAKADFIQREGKYARDMDEVLHAESG
                     HMPEFVERPADYWDAADLYERANGRLFKEVEFALPVELTLDQQKALASEFAQHLTGAER
                     LPYTLAIHAGGGENPHCHLMISERINDGIERPAAQWFKRYNGKTPEKGGAQKTEALKPK
                     AWLEQTREAWADHANRALERAGHDARIDHRTLEAQGIERLPGVHLGPNVVEMEGRGIRT
                     DRADVALNIDTANAQIIDLQEYREAIDHERNRQSEEIQRHQRVSGADRTAGPEHGDTGR
                     RSPAGHEPDQAGQRGAGGGVAESPAPDRGGMGGAGQRVAGGSRRGEQRRAERPERVAGV
                     ALEAMANRDAGFHDAYGGAADRIVALARPDATDNRGRLDLAALGGPMKNDRTLQAIGRQ
                     LKAMGCERFDIGVRDAPTGQMMNREWSAAEVLQNTPWLKRMNAQGNDVYIRPAEQERHG
                     LVLVDDLSEFDLDDMKAEGREPALVVETSPKNYQAWVKVADAAGGELRGQIARTLASEY
                     DADPASADSRHYGRLAGFTNRKDKHTTRAGYQPWVLLRESKGKTATAGPALVQQAGQQI
                     EQAQRQQEKARRLASLELPERQLSRHRRTALDEYRSEMAGLVKRFGHDLSKCDFIAAQK
                     LASRGRSAEEIGKAMAEASPALAERKPGHEADYIERTVSKVMGLPSVQLARAELARAPA
                     PRQRGMDRGGPDFSM"
     CDS             complement(3835..4806)
                     /codon_start=1
                     /product="repB"
                     /label=repB
                     /note="repA-RSF1010; replication gene; derived from 
                     pRL1383a in INSD accession AF403426, a derivative of 
                     RSF1010 INSD accession M28829"
                     /protein_id="AII99631.1"
                     /translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDAPTGQMMNREWSAAEVL
                     QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
                     YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
                     PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELPERQLSRHRRTALD
                     EYRSEMAGLVKRFGHDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
     misc_feature    4735..4740
                     /label=EcoRV
                     /note="EcoRV"
     CDS             complement(4803..5216)
                     /codon_start=1
                     /product="mobB"
                     /label=mobB
                     /note="mobilization gene; derived from pRL1383a in INSD 
                     accession AF403426, a derivative of RSF1010 INSD accession 
                     M28829"
                     /protein_id="AII99625.1"
                     /translation="MNAIDRVKKSRGINELAEQIEPLAQSMATLADEARQVMSQTKQAS
                     EAQAAEWLKAQRQTGAAWVELAKELREVAAEVSSAAQSARSASRGWHWKLWLTVMLASM
                     MPTVVLLIASLLLLDLTPLTTEDGSIWLRLVAR"
     misc_feature    5662..5667
                     /label=AgeI
                     /note="AgeI"
     oriT            6045..6132
                     /note="oriT; origin of transfer from conjugation; from 
                     pRL1383a in INSD accession AF403426, a derivative of 
                     RSF1010 INSD accession M288298"
     misc_feature    6063..6068
                     /label=AgeI
                     /note="AgeI"
     regulatory      6099..6130
                     /regulatory_class="promoter"
                     /note="P2; derived from pRL1383a in INSD accession 
                     AF403426, a derivative of RSF1010 INSD accession M28829; 
                     the P2 promoter is controlled by MobC and MobA provide 
                     transcription of mobC"
     regulatory      complement(6122..6152)
                     /regulatory_class="promoter"
                     /note="P3; derived from pRL1383a in INSD accession 
                     AF403426, a derivative of RSF1010 INSD accession M28829; 
                     the P3 is controlled by MobC and MobA provide transcription
                     of mobA/repB, mobB, and, probably, E-F-repA-repC-operon"
     regulatory      complement(6141..6172)
                     /regulatory_class="promoter"
                     /note="P1; derived from pRL1383a in INSD accession 
                     AF403426, a derivative of RSF1010 INSD accession M28829; 
                     the P1 promoter is controlled by MobC and MobA provide 
                     transcription of mobA/repB, mobB, and, probably, 
                     E-F-repA-repC-operon"
     misc_feature    6163..6447
                     /note="mobilization gene; from pRL1383a in INSD accession 
                     AF403426, a derivative of RSF1010 INSD accession M28829 
                     with the point mutation K14*"
     rep_origin      complement(6380..6879)
                     /direction=LEFT
                     /note="oriV; origin of replication; derived from pRL1383a 
                     in INSD accession AF403426, a derivative of RSF1010 INSD 
                     accession M28829"
     rep_origin      6474..6868
                     /label=RSF1010 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     RSF1010; requires the RSF1010 RepA/B/C proteins for 
                     replication (Scholz et al., 1989)"
     terminator      7035..7078
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     regulatory      complement(7044..7072)
                     /regulatory_class="terminator"
                     /note="TrrnC; rrnC terminator from Lorist6 INSD accession 
                     X98450; Lorist6"
     misc_feature    7111..7116
                     /label=NheI
                     /note="NheI"
     misc_feature    7117..7137
                     /note="GC; GC-adaptor for seamless assembly, flanking BHR 
                     replicons or neutral sites; synthetic"
     misc_feature    7135..7140
                     /label=ZraI
                     /note="ZraI"