LOCUS       62056_9155        7944 bp DNA     circular SYN 14-APR-2019
DEFINITION  Cloning vector pEM008, complete sequence.
ACCESSION   MK431394
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7944)
  AUTHORS   Mancera E, Frazer C, Porman AM, Ruiz-Castro S, Johnson AD, Bennett 
            RJ.
  TITLE     Genetic Modification of Closely Related Candida Species
  JOURNAL   Front Microbiol 10, 357 (2019)
  PUBMED    30941104
REFERENCE   2  (bases 1 to 7944)
  AUTHORS   Mancera E, Frazer C, Porman AM, Ruiz S, Johnson AD, Bennett RJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (23-JAN-2019) Departmento de Ingenieria Genetica, Centro 
            de Investigacion y de Estudios Avanzados del Instituto Politecnico 
            Nacional, Unidad Irapuato, Km. 9.6 Libramiento Norte Carrera 
            Irapuato-Leon, Irapuato, Guanajuato 36824, Mexico
REFERENCE   3  (bases 1 to 7944)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Front 
            Microbiol 10, 357 (2019)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (23-JAN-2019) Departmento de Ingenieria Genetica, Centro de 
            Investigacion y de Estudios Avanzados del Instituto Politecnico 
            Nacional, Unidad Irapuato, Km. 9.6 Libramiento Norte Carrera 
            Irapuato-Leon, Irapuato, Guanajuato 36824, Mexico"
FEATURES             Location/Qualifiers
     source          1..7944
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     22..55
                     /label=FLP recombination target sequence (FRT)
                     /note="FLP recombination target sequence (FRT)"
     misc_feature    62..1048
                     /label=promoter region from Candida tropicalis PCK1
                     /note="promoter region from Candida tropicalis PCK1"
     CDS             1049..2317
                     /label=FLP
                     /note="site-specific recombinase"
     misc_feature    2321..2708
                     /label=terminator region from Candida albicans ACT1
                     /note="terminator region from Candida albicans ACT1"
     misc_feature    2715..3212
                     /label=promoter region from Candida albicans ACT1
                     /note="promoter region from Candida albicans ACT1"
     gene            3213..4439
                     /gene="SAT1"
                     /label=SAT1
     CDS             join(3213..3222,3877..4439)
                     /codon_start=1
                     /transl_table=11
                     /gene="SAT1"
                     /product="streptomycin acetyltransferase"
                     /label=SAT1
                     /note="nourseothricin resistance marker"
                     /protein_id="QBY25788.1"
                     /translation="MDGEEVAALVIDNGSHMKISVIPEQVAETLDAENHFIVREVFDVH
                     LSDQGFELSTRSVSPYRKDYISDDDSDEDSACYGAFIDQELVGKIELNSTWNDLASIEH
                     IVVSHTHRGKGVAHSLIEFAKKWALSRQLLGIRLETQTNNVPACNLYAKCGFTLGGIDL
                     FTYKTRPQVSNETAMYWYWFSGAQDDA"
     misc_feature    4443..4572
                     /label=terminator region from Candida albicans URA3
                     /note="terminator region from Candida albicans URA3"
     protein_bind    complement(4577..4610)
                     /label=FRT (minimal)
                     /note="supports FLP-mediated excision but not integration
                     (Turan and Bode, 2011)"
     primer_bind     complement(4612..4628)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(4665..4683)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4704..4720)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4728..4744)
                     /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(4752..4782)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4797..4818)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(5106..5694)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        5914..6016
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     CDS             6017..6673
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     promoter        complement(7164..7269)
                     /label=AmpR promoter
     rep_origin      7296..7751
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     7892..7908
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        7918..7936
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggtaccgggc cccccctcga ggaagttcct atactttcta gagaatagga acttcggatc
       61 cgtgtgtgta ttctgtataa tttgtaaact ttattaatgt ggtttttttg cttttaaagt
      121 attttgatta cgaaatgacg cgaaatttcc atgaaaaaaa aaagaagaaa gtttgggggt
      181 tcaattttgt acgataacag tgttttttca ctttttccaa ttcccctccc ccgggaatga
      241 agtgacaaac agttgttatt aataactata tatccgaatt aaacactatt tcctgccagg
      301 tgatgaatat actttatggt taccaagttt caaaaaggaa tattggtggg ttagtcggtt
      361 tacaagtttg aaaatcgata ataatcaagt aaaaattatc tttcaaaatc agcaagagaa
      421 gaatatcccc taaaattcaa taaacaattg ttaaataaac aggcaattct aggcaaaatg
      481 cagaatgatg ttaaaaggtt tatgccgcct ttaactcata ctcacatgac ctttttcttc
      541 tgataattta attcaactaa actaagcaaa aaaaaattaa agcacaggaa aggagttaat
      601 taatttttct aaacaaaaca tgttattgtg aataatacac tcaacatatt cattgattgg
      661 ctggcatgcc cagcatcaaa gaaagaaata aaccaaaaga aactccagag tgaaaaatta
      721 atgtgcacta aaatattaaa atccacctcc ttttccccgc gcgcacacaa agcgtagagc
      781 taactaagag aaaaaaacat tcgcctttac cccgggctaa cgtttgtgcc ggtaggtatc
      841 gggatcatgt tcctaataat tcctatgcat ggtattttta ccccagattt caaaagcaca
      901 taaaagcact tccggagaaa agggatagct gctggcccct ataaaagtgg acgaaaattc
      961 cttccaaaaa aaatattttg ttctccttct tttcttcctt tttatcatca ttaacttaat
     1021 aacaaccaag tttaaataat tattcaatat gccacaattt gatatattat gtaaaacacc
     1081 acctaaggtg cttgttcgtc agtttgtgga aaggtttgaa agaccttcag gtgagaaaat
     1141 agcattatgt gctgctgaac taacctattt atgttggatg attacacata acggaacagc
     1201 aatcaagaga gccacattca tgagctataa tactatcata agcaattcgt tgagtttcga
     1261 tattgtcaat aaatcactcc agtttaaata caagacgcaa aaagcaacaa ttttggaagc
     1321 ctcattaaag aaattgattc ctgcttggga atttacaatt attccttact atggacaaaa
     1381 acatcaatct gatatcactg atattgtaag tagtttgcaa ttacagttcg aatcatcgga
     1441 agaagcagat aagggaaata gccacagtaa aaaaatgctt aaagcacttc taagtgaggg
     1501 tgaaagcatc tgggagatca ctgagaaaat actaaattcg tttgagtata cttcgagatt
     1561 tacaaaaaca aaaactttat accaattcct cttcctagct actttcatca attgtggaag
     1621 attcagcgat attaagaacg ttgatccgaa atcatttaaa ttagtccaaa ataagtattt
     1681 gggagtaata atccagtgtt tagtgacaga gacaaagaca agcgttagta ggcacatata
     1741 cttctttagc gcaaggggta ggatcgatcc acttgtatat ttggatgaat ttttgaggaa
     1801 ttctgaacca gtcctaaaac gagtaaatag gaccggcaat tcttcaagca ataaacagga
     1861 ataccaatta ttaaaagata acttagtcag atcgtacaat aaagctttga agaaaaatgc
     1921 gccttattca atctttgcta taaaaaatgg cccaaaatct cacattggaa gacatttgat
     1981 gacctcattt ctttcaatga agggcctaac ggagttgact aatgttgtgg gaaattggag
     2041 cgataagcgt gcttctgccg tggccaggac aacgtatact catcagataa cagcaatacc
     2101 tgatcactac ttcgcactag tttctcggta ctatgcatat gatccaatat caaaggaaat
     2161 gatagcattg aaggatgaga ctaatccaat tgaggagtgg cagcatatag aacagctaaa
     2221 gggtagtgct gaaggaagca tacgataccc cgcatggaat gggataatat cacaggaggt
     2281 actagactac ctttcatcct acataaatag acgcatataa gagtgaaatt ctggaaatct
     2341 ggaaatctgg ttttgtattc ttgttattct tctttttgtt attacatata taacttgtta
     2401 cttttttaaa aaaatctttg tatattttat aaatatataa aactaaattt aagaaaaaga
     2461 gaaaaatgtt ttatttgaga gattgaaatt ttacttgaat ttagcttagc ttttataaag
     2521 tattattatg taaaaaaaca aaacaaatat acattaaaaa gttaagacta taaaatagcc
     2581 acccaaggca tttctatatc ttgttgttgt tgttttcatc ttctgtatca gaggaactta
     2641 ttttattatt ttcgtcacgg gtattttctc ttgtttgatg attcatccca ttcattccat
     2701 cataaaatgt cgagcgtcaa aactagagaa taataaagaa aacgatcttt tcaaaaagaa
     2761 aaaacctttt agttttcctt tgttgttgtt gtgggtgtgt gctatttata ttatatagtt
     2821 tactcataat accataaaat attcggtttg attaggttat tttaataagc taatttgttt
     2881 ctaatcgtgt aatttatgct gtgtatatta agtagtgtgt gcactgccca aaaatgtttg
     2941 ttgtttatag tcggttaaag agaaaaaaga aaaaaagatc catacacaca cgttaattag
     3001 ttgttcaacg taatacactc atattttgtt cttatttgct ttcggtcgct gttctcacca
     3061 agatttattg ccaacgaaac aatttttttt tatatatttt cagatttttc tttttttcct
     3121 ttcctttcct tttctaattt tcactcctgg ttttctttct ttcttagaaa cattatctcg
     3181 atattaatat taaaaaaata taatcattca aaatggacgg tggtatgttt tagtttagct
     3241 tcaattctaa ttgattgatt aatcagttga ttggtttcaa tatgacaaat gggtagggtg
     3301 ggaaaacttc attttcaatt cagatcaaac ttttttgttg tcgacataat atttctcgtt
     3361 tgggatgtta ctgtcacatt aataatacac acacatcagc ttataatttt gaaagtaatt
     3421 tatcagatat gttgtgacga tcaatggaaa tggctaactt caatgtatct gttcttcccc
     3481 tttttcaaag ttcacgtttt ttgattgatt gattgatctg tcggcagtgg tttcaaaacc
     3541 attcggtgag taatcctatc aatcaatgtt acgacaaaag gctcaatatt caaaattgca
     3601 atgttttatg ttttcctacg tgtacttgtg caaggcaatt gattcaacat tgcttttggt
     3661 gtttgacgag tttctagttt ggacttgtgt tgttatctgg gctatacaga tttcccggct
     3721 cactatgaat tttttttttc gacgctcagt gcacacaact ataaacaaca caaacacaaa
     3781 cacagcaaga aaaaaaaaaa acgaacattg aattgaaacc aagccaactg aaaaattcct
     3841 tatttaaatg actgtcatac taacccattt ttatagaaga agttgctgct ttagttatcg
     3901 ataacggttc tcatatgaaa atttcggtga tccctgagca ggtggcggaa acattggatg
     3961 ctgagaacca tttcattgtt cgtgaagtgt tcgatgtgca cctatccgac caaggctttg
     4021 aactatctac cagaagtgtg agcccctacc ggaaggatta catctcggat gatgactctg
     4081 atgaagactc tgcttgctat ggcgcattca tcgaccaaga gcttgtcggg aagattgaac
     4141 tcaactcaac atggaacgat ctagcctcta tcgaacacat tgttgtgtcg cacacgcacc
     4201 gaggcaaagg agtcgcgcac agtctcatcg aatttgcgaa aaagtgggca ctaagcagac
     4261 agctccttgg catacgatta gagacacaaa cgaacaatgt acctgcctgc aatttgtacg
     4321 caaaatgtgg ctttactctc ggcggcattg acctcttcac gtataaaact agacctcaag
     4381 tctcgaacga aacagcgatg tactggtact ggttctcggg agcacaggat gacgcctaac
     4441 atatgtgaag tgtgaagggg gagattttca ctttattaga tttgtatata tgtataataa
     4501 ataaataaat aagttaaata aataattaga taagggtggt aattattact atttacaatc
     4561 aaaggtggtc ctgcaggaag ttcctatact ttctagagaa taggaacttc agatccacta
     4621 gttctagagc ggccgccacc gcggtggagc tccagctttt gttcccttta gtgagggtta
     4681 attgcgcgct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     4741 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     4801 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     4861 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     4921 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     4981 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     5041 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     5101 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     5161 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     5221 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     5281 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     5341 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     5401 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     5461 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     5521 tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct gctgaagcca
     5581 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     5641 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     5701 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     5761 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttcgaccg aataaatacc
     5821 tgtgacggaa gatcacttcg cagaataaat aaatcctggt gtccctgttg ataccgggaa
     5881 gccctgggcc aacttttggc gaaaatgaga cgttgatcgg cacgtaagag gttccaactt
     5941 tcaccataat gaaataagat cactaccggg cgtatttttt gagttgtcga gattttcagg
     6001 agctaaggaa gctaaaatgg agaaaaaaat cactggatat accaccgttg atatatccca
     6061 atggcatcgt aaagaacatt ttgaggcatt tcagtcagtt gctcaatgta cctataacca
     6121 gaccgttcag ctggatatta cggccttttt aaagaccgta aagaaaaata agcacaagtt
     6181 ttatccggcc tttattcaca ttcttgcccg cctgatgaat gctcatccgg aattacgtat
     6241 ggcaatgaaa gacggtgagc tggtgatatg ggatagtgtt cacccttgtt acaccgtttt
     6301 ccatgagcaa actgaaacgt tttcatcgct ctggagtgaa taccacgacg atttccggca
     6361 gtttctacac atatattcgc aagatgtggc gtgttacggt gaaaacctgg cctatttccc
     6421 taaagggttt attgagaata tgtttttcgt ctcagccaat ccctgggtga gtttcaccag
     6481 ttttgattta aacgtggcca atatggacaa cttcttcgcc cccgttttca ccatgggcaa
     6541 atattatacg caaggcgaca aggtgctgat gccgctggcg attcaggttc atcatgccgt
     6601 ttgtgatggc ttccatgtcg gcagaatgct taatgaatta caacagtact gcgatgagtg
     6661 gcagggcggg gcgtaatttt tttaaggcag ttattggtgc ccttaaacgc ctggttgcta
     6721 cgcctgaata agtgataata agcggatgaa tggcagaaat tcgaaagcaa attcgacccg
     6781 gtcgtcggtt cagggcaggg tcgttaaata gccgcttatg tctattgctg gtttaccggt
     6841 ttattgacta ccggaagcag tgtgaccgtg tgcttctcaa atgcctgagg ccagtttgct
     6901 caggctctcc ccgtggaggt aataattgac gatatgatcc tttttttctg atcaaaagtg
     6961 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7021 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7081 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     7141 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcaa
     7201 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     7261 ggttccgcgc acatttcccc gaaaagtgcc acctgacgcg ccctgtagcg gcgcattaag
     7321 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     7381 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     7441 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     7501 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     7561 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     7621 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     7681 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     7741 gcttacaatt tccattcgcc attcaggctg cgcaactgtt gggaagggcg atcggtgcgg
     7801 gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg attaagttgg
     7861 gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga gcgcgcgtaa
     7921 tacgactcac tatagggcga attg
//
