LOCUS       Exported                6689 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pKOS6b, complete sequence.
ACCESSION   JX885593
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6689)
  AUTHORS   Kostner D, Peters B, Mientus M, Liebl W, Ehrenreich A.
  TITLE     Importance of codB for new codA-based markerless gene deletion in 
            Gluconobacter strains
  JOURNAL   Appl. Microbiol. Biotechnol. 97 (18), 8341-8349 (2013)
  PUBMED    23955475
REFERENCE   2  (bases 1 to 6689)
  AUTHORS   Kostner D, Liebl W, Ehrenreich A.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-SEP-2012) Department of Microbiology, TU Munich, Emil 
            Ramannstr. 4, Freising, Bayern 85354, Germany
REFERENCE   3  (bases 1 to 6689)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6689
                     /organism="Cloning vector pKOS6b"
                     /mol_type="other DNA"
                     /db_xref="taxon:1267391"
     CDS             401..1195
                     /codon_start=1
                     /product="kanamycin resistance protein"
                     /label=kanamycin resistance protein
                     /protein_id="AFZ75333.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             1495..2754
                     /codon_start=1
                     /gene="codB"
                     /product="cytosine transporter"
                     /label=codB
                     /protein_id="AFZ75332.1"
                     /translation="MSQDNNFSQGPVPQSARKGVLALTFVMLGLTFFSASMWTGGTLGT
                     GLSYHDFFLAVLIGNLLLGIYTSFLGYIGAKTGLTTHLLARFSFGVKGSWLPSLLLGGT
                     QVGWFGVGVAMFAIPVGKATGLDINLLIAVSGLLMTVTVFFGISALTVLSVIAVPAIAC
                     LGGYSVWLAVNGMGGLDALKAVVPAQPLDFNVALALVVGSFISAGTLTADFVRFGRNAK
                     LAVLVAMVAFFLGNSLMFIFGAAGAAALGMADISDVMIAQGLLLPAIVVLGLNIWTTND
                     NALYASGLGFANITGMSSKTLSVINGIIGTVCALWLYNNFVGWLTFLSAAIPPVGGVII
                     ADYLMNRRRYEHFATTRMMSVNWVAILAVALGIAAGHWLPGIVPVNAVLGGALSYLILN
                     PILNRKTTAAMTHVEANSVE"
     gene            1495..2754
                     /gene="codB"
                     /label=codB
     CDS             2744..4027
                     /codon_start=1
                     /gene="codA"
                     /product="cytosine deaminase"
                     /EC_number="3.5.4.1"
                     /label=codA
                     /protein_id="AFZ75331.1"
                     /translation="MSNNALQTIINARLPGEEGLWQIHLQDGKISAIDAQSGVMPITEN
                     SLDAEQGLVIPPFVEPHIHLDTTQTAGQPNWNQSGTLFEGIERWAERKALLTHDDVKQR
                     AWQTLKWQIANGIQHVRTHVDVSDATLTALKAMLEVKQEVAPWIDLQIVAFPQEGILSY
                     PNGEALLEEALRLGADVVGAIPHFEFTREYGVESLHKTFALAQKYDRLIDVHCDEIDDE
                     QSRFVETVAALAHHEGMGARVTASHTTAMHSYNGAYTSRLFRLLKMSGINFVANPLVNI
                     HLQGRFDTYPKRRGITRVKEMLESGINVCFGHDDVFDPWYPLGTANMLQVLHMGLHVCQ
                     LMGYGQINDGLNLITHHSARTLNLQDYGIAAGNSANLIILPAENGFDALRRQVPVRYSV
                     RGGKVIASTQPAQTTVYLEQPEAIDYKR"
     gene            2744..4027
                     /gene="codA"
                     /label=codA
     CDS             2747..4027
                     /codon_start=1
                     /gene="codA"
                     /product="E. coli cytosine deaminase"
                     /label=codA
                     /note="confers sensitivity to 5-fluorocytosine"
                     /translation="SNNALQTIINARLPGEEGLWQIHLQDGKISAIDAQSGVMPITENS
                     LDAEQGLVIPPFVEPHIHLDTTQTAGQPNWNQSGTLFEGIERWAERKALLTHDDVKQRA
                     WQTLKWQIANGIQHVRTHVDVSDATLTALKAMLEVKQEVAPWIDLQIVAFPQEGILSYP
                     NGEALLEEALRLGADVVGAIPHFEFTREYGVESLHKTFALAQKYDRLIDVHCDEIDDEQ
                     SRFVETVAALAHHEGMGARVTASHTTAMHSYNGAYTSRLFRLLKMSGINFVANPLVNIH
                     LQGRFDTYPKRRGITRVKEMLESGINVCFGHDDVFDPWYPLGTANMLQVLHMGLHVCQL
                     MGYGQINDGLNLITHHSARTLNLQDYGIAAGNSANLIILPAENGFDALRRQVPVRYSVR
                     GGKVIASTQPAQTTVYLEQPEAIDYKR"
     oriT            4432..4541
                     /note="incP origin of transfer"
     promoter        complement(4776..4866)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     promoter        complement(5060..5150)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      5510..6098
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    6386..6407
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6422..6452
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6460..6476
                     /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)."
     primer_bind     6484..6500
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    6509..6565
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(6569..6585)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"