LOCUS       Exported                6065 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Fungal transformation vector pKT300, complete sequence.
ACCESSION   MH286798
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6065)
  AUTHORS   Twaruschek K, Spoerhase P, Michlmayr H, Wiesenberger G, Adam G.
  TITLE     New plasmids allowing positive-negative selection for fungal 
            transformation and efficient Cre-loxP mediated marker recycling
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6065)
  AUTHORS   Twaruschek K, Spoerhase P, Michlmayr H, Wiesenberger G, Adam G.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-MAY-2018) Department of Applied Genetics and Cell 
            Biology, University of Natural Resources and Life Sciences Vienna 
            (BOKU), Konrad Lorenz Strasse 24, Tulln 3430, Austria
REFERENCE   3  (bases 1 to 6065)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6065
                     /organism="Fungal transformation vector pKT300"
                     /mol_type="other DNA"
                     /db_xref="taxon:2211124"
     primer_bind     complement(1..17)
                     /label=SP6 (for sequencing of downstream insert)
                     /note="SP6 (for sequencing of downstream insert)"
     promoter        363..467
                     /gene="bla"
                     /label=AmpR promoter
     CDS             468..1328
                     /codon_start=1
                     /gene="ampR"
                     /product="beta-lactamase"
                     /label=ampR
                     /note="ampicillin resistance protein"
                     /protein_id="AWN56775.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            468..1328
                     /gene="ampR"
                     /label=ampR
     rep_origin      1423..2105
                     /label=ColE1 origin
                     /note="ColE1 origin"
     rep_origin      1499..2087
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2345..2364
                     /label=T7 (for sequencing of upstream insert)
                     /note="T7 (for sequencing of upstream insert)"
     promoter        2345..2363
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_recomb     2434..2467
                     /label=loxP
                     /note="loxP"
     protein_bind    complement(2434..2467)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     regulatory      2481..3227
                     /regulatory_class="promoter"
                     /note="pki; Trichoderma reesei pyruvate kinase I promoter; 
                     accession CP016233"
     regulatory      2634..2643
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             3233..4360
                     /codon_start=1
                     /gene="HSVtk"
                     /product="HSV thymidine kinase"
                     /label=HSVtk
                     /note="no stop codon due to C-terminal resistance gene 
                     fusion"
                     /protein_id="AWN56776.1"
                     /translation="MASYPCHQHASAFDQAARSRGHNNRRTALRPRRQQKATEVRLEQK
                     MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWRVLGASETIANIYTTQ
                     HRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGSSHAPPPALTLIFDR
                     HPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGALPEDRHIDRLAKRQR
                     PGERLDLAMLAAIRRVYGLLANTVRYLQGGGSWREDWGQLSGAAVPPQGAEPQSNAGPR
                     PHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRPMHVFILDYDQSPAGCRDAL
                     LQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     gene            3233..4360
                     /gene="HSVtk"
                     /label=HSVtk
     CDS             4400..5194
                     /codon_start=1
                     /gene="nptII"
                     /product="neomycin phosphotransferase II"
                     /label=nptII
                     /note="kanamycin and G418 resistance protein"
                     /protein_id="AWN56777.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            4400..5194
                     /gene="nptII"
                     /label=nptII
     regulatory      5213..5942
                     /regulatory_class="terminator"
                     /note="cbh2; Trichoderma reesei cellobiohydrolase II 
                     terminator; accession CP016238"
     protein_bind    complement(5957..5990)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        complement(6064..17)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"