LOCUS       40924_26849        9091 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pKLAC1, complete sequence.
ACCESSION   .
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)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 9091)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Appl. 
            Environ. Microbiol."; date: "2005"; volume: "71"; issue: "11"; 
            pages: "7092-7098"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (18-MAR-2005) Research Department, New England Biolabs, 32 Tozer 
            Road, Beverly, MA 01915, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9091
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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"
     promoter        974..1676
                     /label=ADH1 promoter
                     /note="promoter for alcohol dehydrogenase 1"
     CDS             1697..3340
                     /label=amdS
                     /note="acetamidase from Aspergillus nidulans"
     promoter        3979..4635
                     /label=LAC4 promoter (5')
                     /note="5' portion of the Kluyveromyces lactis LAC4
                     promoter"
     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    complement(4708..4724)
                     /label=lac operator
                     /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    complement(4777..4798)
                     /label=CAP binding site
                     /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"
     CDS             complement(5848..6705)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(6706..6810)
                     /label=AmpR promoter
     rep_origin      6837..7292
                     /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"
     promoter        7463..8991
                     /label=LAC4 promoter (3')
                     /note="3' portion of a modified Kluyveromyces lactis LAC4 
                     promoter"
     protein_bind    8997..9013
                     /label=lac operator
                     /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)."
ORIGIN
        1 aagcttgaaa aaaatgaaat tctctactat attagccgca tctactgctt taatttccgt
       61 tgttatggct gctccagttt ctaccgaaac tgacatcgac gatcttccaa tatcggttcc
      121 agaagaagcc ttgattggat tcattgactt aaccggggat gaagtttcct tgttgcctgt
      181 taataacgga acccacactg gtattctatt cttaaacacc accatcgctg aagctgcttt
      241 cgctgacaag gatgatctcg agaaaagaga ggctgaagct agaagagcta gatctcctag
      301 gggtaccgtc gacggcgcgc ctgcggccgc ttaattaagg ccttgaatcg agaatttata
      361 cttagataag tatgtactta caggtatatt tctatgagat actgatgtat acatgcatga
      421 taatatttaa acggttatta gtgccgattg tcttgtgcga taatgacgtt cctatcaaag
      481 caatacactt accacctatt acatgggcca agaaaatatt ttcgaacttg tttagaatat
      541 tagcacagag tatatgatga tatccgttag attatgcatg attcattcct acaacttttt
      601 cgtagcataa ggattaatta cttggatgcc aataaaaaaa aaaaacatcg agaaaatttc
      661 agcatgctca gaaacaattg cagtgtatca aagtaaaaaa aagattttca ctacatgttc
      721 cttttgaaga aagaaaatca tggaacatta gatttacaaa aatttaacca ccgctgatta
      781 acgattagac cgttaagcgc acaacaggtt attagtacag agaaagcatt ctgtggtgtt
      841 gccccggact ttcttttgcg acataggtaa atcgaatacc atcatactat cttttccaat
      901 gactccctaa agaaagactc ttcttcgatg ttgtatacgt tggagcatag ggcaagaatt
      961 gtggcttgag atcatccttt tgttgtttcc gggtgtacaa tatggacttc ctcttttctg
     1021 gcaaccaaac ccatacatcg ggattcctat aataccttcg ttggtctccc taacatgtag
     1081 gtggcggagg ggagatatac aatagaacag ataccagaca agacataatg ggctaaacaa
     1141 gactacacca attacactgc ctcattgatg gtggtacata acgaactaat actgtagccc
     1201 tagacttgat agccatcatc atatcgaagt ttcactaccc tttttccatt tgccatctat
     1261 tgaagtaata ataggcgcat gcaacttctt ttcttttttt ttcttttctc tctcccccgt
     1321 tgttgtctca ccatatccgc aatgacaaaa aaatgatgga agacactaaa ggaaaaaatt
     1381 aacgacaaag acagcaccaa cagatgtcgt tgttccagag ctgatgaggg gtatctcgaa
     1441 gcacacgaaa ctttttcctt ccttcattca cgcacactac tctctaatga gcaacggtat
     1501 acggccttcc ttccagttac ttgaatttga aataaaaaaa agtttgctgt cttgctatca
     1561 agtataaata gacctgcaat tattaatctt ttgtttcctc gtcattgttc tcgttccctt
     1621 tcttccttgt ttctttttct gcacaatatt tcaagctata ccaagcatac aatcaaggaa
     1681 ttccggatcc gccaccatgc ctcaatcctg ggaagaactg gccgctgata agcgcgcccg
     1741 cctcgcaaaa accatccctg atgaatggaa agtccagacg ctgcctgcgg aagacagcgt
     1801 tattgatttc ccaaagaaat cggggatcct ttcagaggcc gaactgaaga tcacagaggc
     1861 ttccgctgcg gatcttgtgt ccaagctggc ggccggagag ttgacctcgg tggaagttac
     1921 gctagcattc tgtaaacggg cagcaatcgc ccagcagtta acaaactgcg cccacgagtt
     1981 cttccctgac gccgctctcg cgcaggcaag ggaactcgat gaatactacg caaagcacaa
     2041 gagacccgtt ggtccactcc atggcctccc catctctctc aaagaccagc ttcgagtcaa
     2101 gggctacgaa acatcaatgg gctacatctc atggctaaac aagtacgacg aaggggactc
     2161 ggttctgaca accatgctcc gcaaagccgg tgccgtcttc tacgtcaaga cctctgtccc
     2221 gcagaccctg atggtctgcg agacagtcaa caacatcatc gggcgcaccg tcaacccacg
     2281 caacaagaac tggtcgtgcg gcggcagttc tggtggtgag ggtgcgatcg ttgggattcg
     2341 tggtggcgtc atcggtgtag gaacggatat cggtggctcg attcgagtgc cggccgcgtt
     2401 caacttcctg tacggtctaa ggccgagtca tgggcggctg ccgtatgcaa agatggcgaa
     2461 cagcatggag ggtcaggaga cggtgcacag cgttgtcggg ccgattacgc actctgttga
     2521 ggacctccgc ctcttcacca aatccgtcct cggtcaggag ccatggaaat acgactccaa
     2581 ggtcatcccc atgccctggc gccagtccga gtcggacatt attgcctcca agatcaagaa
     2641 cggcgggctc aatatcggct actacaactt cgacggcaat gtccttccac accctcctat
     2701 cctgcgcggc gtggaaacta ccgtcgccgc actcgccaaa gccggtcaca ccgtgacccc
     2761 gtggacgcca tacaagcacg atttcggcca cgatctcatc tcccatatct acgcggctga
     2821 cggcagcgcc gacgtaatgc gcgatatcag tgcatccggc gagccggcga ttccaaatat
     2881 caaagaccta ctgaacccga acatcaaagc tgttaacatg aacgagctct gggacacgca
     2941 tctccagaag tggaattacc agatggagta ccttgagaaa tggcgggagg ctgaagaaaa
     3001 ggccgggaag gaactggacg ccatcatcgc gccgattacg cctaccgctg cggtacggca
     3061 tgaccagttc cggtactatg ggtatgcctc tgtgatcaac ctgctggatt tcacgagcgt
     3121 ggttgttccg gttacctttg cggataagaa catcgataag aagaatgaga gtttcaaggc
     3181 ggttagtgag cttgatgccc tcgtgcagga agagtatgat ccggaggcgt accatggggc
     3241 accggttgca gtgcaggtta tcggacggag actcagtgaa gagaggacgt tggcgattgc
     3301 agaggaagtg gggaagttgc tgggaaatgt ggtgactcca tagcccgggg ggggctcgat
     3361 cccctcgcga gttggttcag ctgctgcctg aggctggacg acctcgcgga gttctaccgg
     3421 cagtgcaaat ccgtcggcat ccaggaaacc agcagcggct atccgcgcat ccatgccccc
     3481 gaactgcagg agtggggagg cacgatggcc gctttggtcg atctagatta cgtggaagaa
     3541 aggtagtaaa agtagtagta taagtagtaa aaagaggtaa aaagagaaaa ccggctacat
     3601 actagagaag cacgtacaca aaaactcata ggcacttcat catacgacag tttcttgatg
     3661 cattataata gtgtattaga tattttcaga aatatgcata gaacctcctc ttgcctttac
     3721 tttttataca tagaacattg gcagatttac ttacactact ttgtttctac gccatttctt
     3781 ttgttttcaa cacttagaca agttgttgag aaccggacta ctaaaaagca atgttcccac
     3841 tgaaaatcat gtacctgcag gataataacc ccctaattct gcatcgatcc agtatgtttt
     3901 tttttctcta ctcattttta cctgaagata gagcttctaa aacaaaaaaa atcagtgatt
     3961 acatgcatat tgtgtgttct agtaaccaaa ggaaaggaac agatagataa aattccgaga
     4021 ctgtcaaatt aggttttttt cttttttttt ggcgggagtc agtgggccga aatatgttct
     4081 tggcctagaa cttaatctgg tttgatcatg ccaatacttg cctgagtgcc cgactttttg
     4141 cccaccctct tgccttctgt catccttcaa aacccacctg ttttccagcc gtatcttcgc
     4201 tcgcatctac acatactgtg ccatatcttg tgtgtagccg gacgtgacta tgaccaaaaa
     4261 caaacaagga gaactgttcg ccgatttgta acactcctgc atccatccaa gtgggtatgc
     4321 gctatgcaat gttaagctag gtcaggtcag accaggtcca aggacagcaa cttgactgta
     4381 tgcaaccttt accatctttg cacagaacat acttgtagct agctagttac acttatggac
     4441 cgaaaaggca ccccaccatg tctgtccggc tttagagtac ggccgcagac cgctgatttg
     4501 ccttgccaag cagtagtcac aatgcatcgc atgagcacac gggcacgggc acgggcacag
     4561 gaaccattgg caaaaatacc agatacacta taccgacgta tatcaagccc aagtttaaaa
     4621 ttcctaaatt tccgcggcta cttttcaatt ccctatagtg agtcgtatta aattcgtaat
     4681 catgtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     4741 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     4801 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     4861 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     4921 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     4981 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     5041 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     5101 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     5161 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     5221 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     5281 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     5341 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     5401 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     5461 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     5521 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     5581 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     5641 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     5701 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     5761 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     5821 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     5881 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     5941 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     6001 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     6061 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     6121 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     6181 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     6241 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     6301 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     6361 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     6421 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     6481 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     6541 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     6601 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     6661 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     6721 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     6781 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgc
     6841 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     6901 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     6961 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     7021 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     7081 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     7141 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     7201 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     7261 gaattttaac aaaatattaa cgcttacaat ttccattcgc cattcaggct gcgcaactgt
     7321 tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa agggggatgt
     7381 gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg ttgtaaaacg
     7441 acggccagtg ccaagctccc gcggggatcg actcataaaa tagtaacctt ctaatgcgta
     7501 tctattgact accaaccatt agtgtggttg cagaaggcgg aattctccct tcttcgaatt
     7561 cagcttgctt tttcattttt tattttccat ttttcagttt ttgtttgtgt cgaatttagc
     7621 cagttgcttc tccaagatga aaaaaacccc tgcgcagttt ctgtgctgca agatcctaat
     7681 cgacttttcc accccccaca aaagtaaatg ttcttttgtt acattcgcgt gggtagctag
     7741 ctccccgaat cttcaaagga cttagggact gcactacatc agagtgtgtt cacctggttt
     7801 gctgcctggt ttgaaagaaa agagcaggga actcgcgggt tcccggcgaa taatcatgcg
     7861 atagtccttt ggccttccaa gtcgcatgta gagtagacaa cagacaggga gggcaggaag
     7921 gatctttcac tgagatcctg tatcttgttg ggtaagtcgg atgaaagggg aatcgtatga
     7981 gattggagag gatgcggaag aggtaacgcc ttttgttaac ttgtttaatt attatggggc
     8041 aggcgagagg gggaggaatg tatgtgtgtg aggcgggcga gacggagcca tccaggccag
     8101 gtagaaatag agaaagccga atgttagaca atatggcagc gtagtagagt aggtaggtag
     8161 gcaagtactg ctagcaaaga ggagaagggt aagctcactc ttcgcattcc acaccgttag
     8221 tgtgtcagtt tgaacaaaaa aacaatcatc ataccaattg atggactgtg gactggcttt
     8281 tggaacggct tttcggactg cgattattcg tgaggaatca aggtaggaat ttggtcatat
     8341 ttacggacaa cagtgggtga ttcccatatg gagtaggaaa acgagatcat ggtatcctca
     8401 gatatgttgc ggaattctgt tcaccgcaaa gttcagggtg ctctggtggg tttcggttgg
     8461 tctttgcttt gcttctccct tgtcttgcat gttaataata gcctagcctg tgagccgaaa
     8521 cttagggtag gcttagtgtt ggaacgtaca tatgtatcac gttgacttgg tttaaccagg
     8581 cgacctggta gccagccata cccacacacg ttttttgtat cttcagtata gttgtgaaaa
     8641 gtgtagcgga aatttgtggt ccgagcaaca gcgtcttttt ctagtagtgc ggtcggttac
     8701 ttggttgaca ttggtatttg gactttgttg ctacaccatt cactacttga agtcgagtgt
     8761 gaagggtatg atttctagtg gtgaacacct ttagttacgt aatgttttca ttgctgtttt
     8821 acttgagatt tcgattgaga aaaaggtatt taatagctcg aatcaatgtg agaacagaga
     8881 gaagatgttc ttccctaact cgaaaggtat atgaggcttg tgtttcttag gagaattatt
     8941 attcttttgt tatgttgcgc ttgtagttgg aaaaggtgaa gagacaaaag ctggaattgt
     9001 gagcggataa caagctcaac acttgaaatt taggaaagag cagaatttgg caaaaaaaat
     9061 aaaaaaaaaa taaacacaca tactcatcga g
//
