LOCUS       62056_9150        7557 bp DNA     circular SYN 14-APR-2019
DEFINITION  Cloning vector pEM004, complete sequence.
ACCESSION   MK425748
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7557)
  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 7557)
  AUTHORS   Mancera E, Frazer C, Porman AM, Ruiz S, Johnson AD, Bennett RJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (22-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 7557)
  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 
            (22-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..7557
                     /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..661
                     /label=promoter region from Candida tropicalis MAL2
                     /note="promoter region from Candida tropicalis MAL2"
     CDS             662..1930
                     /label=FLP
                     /note="site-specific recombinase"
     misc_feature    1934..2321
                     /label=terminator region from Candida albicans ACT1
                     /note="terminator region from Candida albicans ACT1"
     misc_feature    2328..2825
                     /label=promoter region from Candida albicans ACT1
                     /note="promoter region from Candida albicans ACT1"
     gene            2826..4052
                     /gene="SAT1"
                     /label=SAT1
     CDS             join(2826..2835,3490..4052)
                     /codon_start=1
                     /transl_table=11
                     /gene="SAT1"
                     /product="streptomycin acetyltransferase"
                     /label=SAT1
                     /note="nourseothricin resistance marker"
                     /protein_id="QBY25891.1"
                     /translation="MDGEEVAALVIDNGSHMKISVIPEQVAETLDAENHFIVREVFDVH
                     LSDQGFELSTRSVSPYRKDYISDDDSDEDSACYGAFIDQELVGKIELNSTWNDLASIEH
                     IVVSHTHRGKGVAHSLIEFAKKWALSRQLLGIRLETQTNNVPACNLYAKCGFTLGGIDL
                     FTYKTRPQVSNETAMYWYWFSGAQDDA"
     misc_feature    4056..4185
                     /label=terminator region from Candida albicans URA3
                     /note="terminator region from Candida albicans URA3"
     protein_bind    complement(4190..4223)
                     /label=FRT (minimal)
                     /note="supports FLP-mediated excision but not integration
                     (Turan and Bode, 2011)"
     primer_bind     complement(4225..4241)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(4278..4296)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4317..4333)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(4341..4357)
                     /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(4365..4395)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(4410..4431)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(4719..5307)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        5527..5629
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     CDS             5630..6286
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     promoter        complement(6777..6882)
                     /label=AmpR promoter
     rep_origin      6909..7364
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     primer_bind     7505..7521
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        7531..7549
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 ggtaccgggc cccccctcga ggaagttcct atactttcta gagaatagga acttcggatc
       61 cgttaaaagc cctgtatacc gaattactaa gaaattattg tctgatttgt aattcagaac
      121 gcaaagtgta atgccctgta tcttattttt ttttttttgc agctatacga taagaatctc
      181 atttttttgg tgccaatgcg atcgtttata cgcaattcct aattgaaacg tctcaacaaa
      241 aaaattgtaa atcatcttca attggtagct ttaataactc taaattcaag ggggggattg
      301 ccagcgtatc tgtataaacg agtgttcttt tctactacat tgctgtggat actattaata
      361 gacaaacagt agtgtgtgta tacacataca gaatccatct ctctccgcca cattgtttcc
      421 aataaaaaaa tattatccag taattggtgt ttttgtgttt ctatggatca attctcggaa
      481 aaattatcgc gagaacaaag agcaagtgtg gggagatggg gagtcaacac agcaagagaa
      541 caaaagcttt aaatagtcca tcttttcaac ggacaagctg aaaaaaaaga gcaattactt
      601 gttactcttc aatttttttt ttctctactt tttatatatt cttcaaacaa aactattaat
      661 tatgccacaa tttgatatat tatgtaaaac accacctaag gtgcttgttc gtcagtttgt
      721 ggaaaggttt gaaagacctt caggtgagaa aatagcatta tgtgctgctg aactaaccta
      781 tttatgttgg atgattacac ataacggaac agcaatcaag agagccacat tcatgagcta
      841 taatactatc ataagcaatt cgttgagttt cgatattgtc aataaatcac tccagtttaa
      901 atacaagacg caaaaagcaa caattttgga agcctcatta aagaaattga ttcctgcttg
      961 ggaatttaca attattcctt actatggaca aaaacatcaa tctgatatca ctgatattgt
     1021 aagtagtttg caattacagt tcgaatcatc ggaagaagca gataagggaa atagccacag
     1081 taaaaaaatg cttaaagcac ttctaagtga gggtgaaagc atctgggaga tcactgagaa
     1141 aatactaaat tcgtttgagt atacttcgag atttacaaaa acaaaaactt tataccaatt
     1201 cctcttccta gctactttca tcaattgtgg aagattcagc gatattaaga acgttgatcc
     1261 gaaatcattt aaattagtcc aaaataagta tttgggagta ataatccagt gtttagtgac
     1321 agagacaaag acaagcgtta gtaggcacat atacttcttt agcgcaaggg gtaggatcga
     1381 tccacttgta tatttggatg aatttttgag gaattctgaa ccagtcctaa aacgagtaaa
     1441 taggaccggc aattcttcaa gcaataaaca ggaataccaa ttattaaaag ataacttagt
     1501 cagatcgtac aataaagctt tgaagaaaaa tgcgccttat tcaatctttg ctataaaaaa
     1561 tggcccaaaa tctcacattg gaagacattt gatgacctca tttctttcaa tgaagggcct
     1621 aacggagttg actaatgttg tgggaaattg gagcgataag cgtgcttctg ccgtggccag
     1681 gacaacgtat actcatcaga taacagcaat acctgatcac tacttcgcac tagtttctcg
     1741 gtactatgca tatgatccaa tatcaaagga aatgatagca ttgaaggatg agactaatcc
     1801 aattgaggag tggcagcata tagaacagct aaagggtagt gctgaaggaa gcatacgata
     1861 ccccgcatgg aatgggataa tatcacagga ggtactagac tacctttcat cctacataaa
     1921 tagacgcata taagagtgaa attctggaaa tctggaaatc tggttttgta ttcttgttat
     1981 tcttcttttt gttattacat atataacttg ttactttttt aaaaaaatct ttgtatattt
     2041 tataaatata taaaactaaa tttaagaaaa agagaaaaat gttttatttg agagattgaa
     2101 attttacttg aatttagctt agcttttata aagtattatt atgtaaaaaa acaaaacaaa
     2161 tatacattaa aaagttaaga ctataaaata gccacccaag gcatttctat atcttgttgt
     2221 tgttgttttc atcttctgta tcagaggaac ttattttatt attttcgtca cgggtatttt
     2281 ctcttgtttg atgattcatc ccattcattc catcataaaa tgtcgagcgt caaaactaga
     2341 gaataataaa gaaaacgatc ttttcaaaaa gaaaaaacct tttagttttc ctttgttgtt
     2401 gttgtgggtg tgtgctattt atattatata gtttactcat aataccataa aatattcggt
     2461 ttgattaggt tattttaata agctaatttg tttctaatcg tgtaatttat gctgtgtata
     2521 ttaagtagtg tgtgcactgc ccaaaaatgt ttgttgttta tagtcggtta aagagaaaaa
     2581 agaaaaaaag atccatacac acacgttaat tagttgttca acgtaataca ctcatatttt
     2641 gttcttattt gctttcggtc gctgttctca ccaagattta ttgccaacga aacaattttt
     2701 ttttatatat tttcagattt ttcttttttt cctttccttt ccttttctaa ttttcactcc
     2761 tggttttctt tctttcttag aaacattatc tcgatattaa tattaaaaaa atataatcat
     2821 tcaaaatgga cggtggtatg ttttagttta gcttcaattc taattgattg attaatcagt
     2881 tgattggttt caatatgaca aatgggtagg gtgggaaaac ttcattttca attcagatca
     2941 aacttttttg ttgtcgacat aatatttctc gtttgggatg ttactgtcac attaataata
     3001 cacacacatc agcttataat tttgaaagta atttatcaga tatgttgtga cgatcaatgg
     3061 aaatggctaa cttcaatgta tctgttcttc ccctttttca aagttcacgt tttttgattg
     3121 attgattgat ctgtcggcag tggtttcaaa accattcggt gagtaatcct atcaatcaat
     3181 gttacgacaa aaggctcaat attcaaaatt gcaatgtttt atgttttcct acgtgtactt
     3241 gtgcaaggca attgattcaa cattgctttt ggtgtttgac gagtttctag tttggacttg
     3301 tgttgttatc tgggctatac agatttcccg gctcactatg aatttttttt ttcgacgctc
     3361 agtgcacaca actataaaca acacaaacac aaacacagca agaaaaaaaa aaaacgaaca
     3421 ttgaattgaa accaagccaa ctgaaaaatt ccttatttaa atgactgtca tactaaccca
     3481 tttttataga agaagttgct gctttagtta tcgataacgg ttctcatatg aaaatttcgg
     3541 tgatccctga gcaggtggcg gaaacattgg atgctgagaa ccatttcatt gttcgtgaag
     3601 tgttcgatgt gcacctatcc gaccaaggct ttgaactatc taccagaagt gtgagcccct
     3661 accggaagga ttacatctcg gatgatgact ctgatgaaga ctctgcttgc tatggcgcat
     3721 tcatcgacca agagcttgtc gggaagattg aactcaactc aacatggaac gatctagcct
     3781 ctatcgaaca cattgttgtg tcgcacacgc accgaggcaa aggagtcgcg cacagtctca
     3841 tcgaatttgc gaaaaagtgg gcactaagca gacagctcct tggcatacga ttagagacac
     3901 aaacgaacaa tgtacctgcc tgcaatttgt acgcaaaatg tggctttact ctcggcggca
     3961 ttgacctctt cacgtataaa actagacctc aagtctcgaa cgaaacagcg atgtactggt
     4021 actggttctc gggagcacag gatgacgcct aacatatgtg aagtgtgaag ggggagattt
     4081 tcactttatt agatttgtat atatgtataa taaataaata aataagttaa ataaataatt
     4141 agataagggt ggtaattatt actatttaca atcaaaggtg gtcctgcagg aagttcctat
     4201 actttctaga gaataggaac ttcagatcca ctagttctag agcggccgcc accgcggtgg
     4261 agctccagct tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca
     4321 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     4381 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     4441 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     4501 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     4561 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4621 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4681 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4741 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4801 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4861 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4921 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4981 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     5041 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     5101 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     5161 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     5221 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     5281 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5341 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5401 ttcacctaga tccttttcga ccgaataaat acctgtgacg gaagatcact tcgcagaata
     5461 aataaatcct ggtgtccctg ttgataccgg gaagccctgg gccaactttt ggcgaaaatg
     5521 agacgttgat cggcacgtaa gaggttccaa ctttcaccat aatgaaataa gatcactacc
     5581 gggcgtattt tttgagttgt cgagattttc aggagctaag gaagctaaaa tggagaaaaa
     5641 aatcactgga tataccaccg ttgatatatc ccaatggcat cgtaaagaac attttgaggc
     5701 atttcagtca gttgctcaat gtacctataa ccagaccgtt cagctggata ttacggcctt
     5761 tttaaagacc gtaaagaaaa ataagcacaa gttttatccg gcctttattc acattcttgc
     5821 ccgcctgatg aatgctcatc cggaattacg tatggcaatg aaagacggtg agctggtgat
     5881 atgggatagt gttcaccctt gttacaccgt tttccatgag caaactgaaa cgttttcatc
     5941 gctctggagt gaataccacg acgatttccg gcagtttcta cacatatatt cgcaagatgt
     6001 ggcgtgttac ggtgaaaacc tggcctattt ccctaaaggg tttattgaga atatgttttt
     6061 cgtctcagcc aatccctggg tgagtttcac cagttttgat ttaaacgtgg ccaatatgga
     6121 caacttcttc gcccccgttt tcaccatggg caaatattat acgcaaggcg acaaggtgct
     6181 gatgccgctg gcgattcagg ttcatcatgc cgtttgtgat ggcttccatg tcggcagaat
     6241 gcttaatgaa ttacaacagt actgcgatga gtggcagggc ggggcgtaat ttttttaagg
     6301 cagttattgg tgcccttaaa cgcctggttg ctacgcctga ataagtgata ataagcggat
     6361 gaatggcaga aattcgaaag caaattcgac ccggtcgtcg gttcagggca gggtcgttaa
     6421 atagccgctt atgtctattg ctggtttacc ggtttattga ctaccggaag cagtgtgacc
     6481 gtgtgcttct caaatgcctg aggccagttt gctcaggctc tccccgtgga ggtaataatt
     6541 gacgatatga tccttttttt ctgatcaaaa gtgctcatca ttggaaaacg ttcttcgggg
     6601 cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc cactcgtgca
     6661 cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc aaaaacagga
     6721 aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat actcatactc
     6781 ttcctttttc aatattattg aagcatttat caagggttat tgtctcatga gcggatacat
     6841 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     6901 gccacctgac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     6961 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     7021 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     7081 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     7141 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     7201 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     7261 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     7321 aaaatttaac gcgaatttta acaaaatatt aacgcttaca atttccattc gccattcagg
     7381 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctggcg
     7441 aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc ccagtcacga
     7501 cgttgtaaaa cgacggccag tgagcgcgcg taatacgact cactataggg cgaattg
//
