LOCUS       Exported                9091 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pKLAC1, complete sequence.
ACCESSION   AY968582
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9091)
  AUTHORS   Colussi PA, Taron CH.
  TITLE     Kluyveromyces lactis LAC4 promoter variants that lack function in 
            bacteria but retain full function in K. lactis
  JOURNAL   Appl. Environ. Microbiol. 71 (11), 7092-7098 (2005)
  PUBMED    16269745
REFERENCE   2  (bases 1 to 9091)
  AUTHORS   Colussi PA, Taron CH.
  TITLE     Direct Submission
  JOURNAL   Submitted (18-MAR-2005) Research Department, New England Biolabs, 32
            Tozer Road, Beverly, MA 01915, USA
REFERENCE   3  (bases 1 to 9091)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..9091
                     /organism="Expression vector pKLAC1"
                     /host="Kluyveromyces lactis"
                     /mol_type="other DNA"
                     /db_xref="taxon:322510"
     misc_feature    14..334
                     /note="Kluyveromyces lactis alpha mating factor pre-pro 
                     secretion leader"
     misc_feature    290..338
                     /label=multiple cloning site
                     /note="multiple cloning site"
     terminator      355..973
                     /label=LAC4 terminator
                     /note="transcription terminator for Kluyveromyces lactis 
                     LAC4"
     regulatory      355..937
                     /regulatory_class="terminator"
                     /note="Kluyveromyces lactis LAC4 transcription terminator"
     regulatory      994..1696
                     /regulatory_class="promoter"
                     /note="Saccharomyces cerevisiae ADH1 promoter"
     CDS             1697..3343
                     /codon_start=1
                     /gene="amdS"
                     /product="acetamidase"
                     /label=amdS
                     /note="Aspergillus nidulans acetamidase"
                     /protein_id="AAX83680.1"
                     /translation="MPQSWEELAADKRARLAKTIPDEWKVQTLPAEDSVIDFPKKSGIL
                     SEAELKITEASAADLVSKLAAGELTSVEVTLAFCKRAAIAQQLTNCAHEFFPDAALAQA
                     RELDEYYAKHKRPVGPLHGLPISLKDQLRVKGYETSMGYISWLNKYDEGDSVLTTMLRK
                     AGAVFYVKTSVPQTLMVCETVNNIIGRTVNPRNKNWSCGGSSGGEGAIVGIRGGVIGVG
                     TDIGGSIRVPAAFNFLYGLRPSHGRLPYAKMANSMEGQETVHSVVGPITHSVEDLRLFT
                     KSVLGQEPWKYDSKVIPMPWRQSESDIIASKIKNGGLNIGYYNFDGNVLPHPPILRGVE
                     TTVAALAKAGHTVTPWTPYKHDFGHDLISHIYAADGSADVMRDISASGEPAIPNIKDLL
                     NPNIKAVNMNELWDTHLQKWNYQMEYLEKWREAEEKAGKELDAIIAPITPTAAVRHDQF
                     RYYGYASVINLLDFTSVVVPVTFADKNIDKKNESFKAVSELDALVQEEYDPEAYHGAPV
                     AVQVIGRRLSEERTLAIAEEVGKLLGNVVTP"
     gene            1697..3343
                     /gene="amdS"
                     /label=amdS
     promoter        3979..4635
                     /label=LAC4 promoter (5)
                     /note="5 portion of the Kluyveromyces lactis LAC4 
                     promoter"
     regulatory      4052..4632
                     /regulatory_class="promoter"
                     /note="modified Kluyveromyces lactis LAC4 promoter 5 end"
     promoter        complement(4652..4670)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4684..4700)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4708..4724
                     /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)."
     promoter        complement(4732..4762)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4777..4798
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(5086..5674)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     gene            complement(5844..6705)
                     /gene="bla"
                     /label=bla
     CDS             complement(5845..6705)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /note="confers ampicillin resistance in Escherichia coli"
                     /protein_id="AAX83681.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6706..6810)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6837..7292
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     7433..7449
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      7459..9091
                     /regulatory_class="promoter"
                     /note="modified Kluyveromyces lactis LAC4 promoter 3 end"
     promoter        7463..8991
                     /label=LAC4 promoter (3)
                     /note="3 portion of a modified Kluyveromyces lactis LAC4 
                     promoter"
     protein_bind    8997..9013
                     /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)."