LOCUS       Exported                3937 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pAK405, complete sequence.
ACCESSION   JQ432562
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3937)
  AUTHORS   Kaczmarczyk A, Vorholt JA, Francez-Charlot A.
  TITLE     Markerless gene deletion system for sphingomonads
  JOURNAL   Appl. Environ. Microbiol. 78 (10), 3774-3777 (2012)
  PUBMED    22427496
REFERENCE   2  (bases 1 to 3937)
  AUTHORS   Kaczmarczyk A, Vorholt JA, Francez-Charlot A.
  TITLE     Direct Submission
  JOURNAL   Submitted (20-JAN-2012) Department of Biology, Institute of 
            Microbiology, ETH Zurich, Wolfgang-Pauli-Str. 10, Zurich, Zurich 
            8093, Switzerland
REFERENCE   3  (bases 1 to 3937)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..3937
                     /organism="Cloning vector pAK405"
                     /mol_type="other DNA"
                     /db_xref="taxon:1166467"
     CDS             346..1140
                     /codon_start=1
                     /gene="nptII"
                     /product="NPTII"
                     /EC_number="2.7.1.95"
                     /label=nptII
                     /note="neomycin phosphotransferase; confers kanamycin 
                     resistance"
                     /protein_id="AFH41828.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            346..1140
                     /gene="nptII"
                     /label=nptII
     regulatory      1167..1207
                     /regulatory_class="promoter"
                     /gene="Psyn2"
                     /note="synthetic bacterial promoter"
     gene            1167..1207
                     /gene="Psyn2"
                     /label=Psyn2
     gene            1221..1605
                     /gene="rpsL1"
                     /label=rpsL1
     regulatory      1221..1229
                     /regulatory_class="ribosome_binding_site"
                     /gene="rpsL1"
     CDS             1234..1605
                     /codon_start=1
                     /gene="rpsL1"
                     /product="30S ribosomal subunit protein S12"
                     /label=rpsL1
                     /note="rpsL1; S12(R88K); from Sphingomonas sp. Fr1 with 
                     R88K amino acid substitution; can be used as a 
                     counterselection marker in sphingomonads"
                     /protein_id="AFH41829.1"
                     /translation="MPTINQLVRKGRDPQKAKSKVPAMEQNPQKRGVCTRVYTTTPKKP
                     NSALRKVAKVRLTNSREVISYIPGEGHNLQEHSVVLIRGGRVKDLPGVRYHVLRGVLDT
                     QGVKDRRQSRSKYGAKRPK"
     oriT            1625..1734
                     /note="incP origin of transfer"
     repeat_region   1672..1770
                     /label=RP4 oriT
                     /rpt_family="RP4 oriT"
                     /rpt_unit_range=1684 .. 1723
                     /note="RP4 oriT"
     promoter        complement(1969..2059)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     promoter        complement(2253..2343)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     rep_origin      2687..3306
                     /label=pBR322 oriV
                     /note="pBR322 oriV"
     rep_origin      2703..3291
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    3579..3600
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3615..3645
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3653..3669
                     /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     3677..3693
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             3694..3924
                     /codon_start=1
                     /gene="lacZ"
                     /product="LacZ"
                     /label=lacZ
                     /note="beta-galactosidase fragment; allows blue/white 
                     screening by alpha complementation"
                     /protein_id="AFH41830.1"
                     /translation="MITNSSSVPGDPLESTCRHASLALAVVLQRRDWENPGVTQLNRLA
                     AHPPFASWRNSEEARTDRPSQQLRSLNGEWR"
     gene            3694..3924
                     /gene="lacZ"
                     /label=lacZ
     misc_feature    3702..3758
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(3762..3778)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"