LOCUS       40924_47208        9353 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pXIPHOS, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9353)
  AUTHORS   Higgins DA, Young MK, Tremaine M, Sardi M, Fletcher JM, Agnew M, Liu
            L, Dickinson Q, Peris D, Wrobel RL, Hittinger CT, Gasch AP, Singer 
            SW, Simmons BA, Landick R, Thelen MP, Sato TK.
  TITLE     Natural Variation in the Multidrug Efflux Pump SGE1 Underlies Ionic 
            Liquid Tolerance in Yeast
  JOURNAL   Genetics 210 (1), 219-234 (2018)
  PUBMED    30045857
REFERENCE   2  (bases 1 to 9353)
  AUTHORS   Wrobel RL, Alexander WG, Hittinger CT.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-FEB-2018) Laboratory of Genetics/Great Lakes Bioenergy
            Research Center, University of Wisconsin-Madison, 425 Henry Mall, 
            4102 Genetics Biotechnology Center, Madison, WI 53711, United States
REFERENCE   3  (bases 1 to 9353)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 9353)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Genetics"; 
            date: "2018"; volume: "210"; issue: "1"; pages: "219-234"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (02-FEB-2018) Laboratory of Genetics/Great Lakes Bioenergy Research 
            Center, University of Wisconsin-Madison, 425 Henry Mall, 4102 
            Genetics Biotechnology Center, Madison, WI 53711, United States"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..9353
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..45
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(79..97)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     gene            164..1283
                     /label=natMX6
                     /note="yeast selectable marker conferring nourseothricin 
                     resistance"
     CDS             2052..6155
                     /codon_start=1
                     /label=Cas9
                     /note="Cas9 (Csn1) endonuclease from the Streptococcus
                     pyogenes Type II CRISPR/Cas system"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     terminator      6192..6381
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     rep_origin      complement(6444..7032)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(7206..8063)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(8064..8168)
                     /label=AmpR promoter
     rep_origin      complement(join(8195..9353,1..2))
                     /direction=LEFT
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
ORIGIN
        1 gtatatgtgt tatgtatgag cgcgcgtaat acgactcact atagggcgaa ttgggtacgc
       61 ggccgccagc ttttgttccc tttagtgagg gttaattgcg cgcttggcgt aggtctagag
      121 atctgtttag cttgcctcgt ccccgccggg tcacccggcc agcgacatgg aggcccagaa
      181 taccctcctt gacagtcttg acgtgcgcag ctcaggggca tgatgtgact gtcgcccgta
      241 catttagccc atacatcccc atgtataatc atttgcatcc atacattttg atggccgcac
      301 ggcgcgaagc aaaaattacg gctcctcgct gcagacctgc gagcagggaa acgctcccct
      361 cacagacgcg ttgaattgtc cccacgccgc gcccctgtag agaaatataa aaggttagga
      421 tttgccactg aggttcttct ttcatatact tccttttaaa atcttgctag gatacagttc
      481 tcacatcaca tccgaacata aacaaccatg ggtaccactc ttgacgacac ggcttaccgg
      541 taccgcacca gtgtcccggg ggacgccgag gccatcgagg cactggatgg gtccttcacc
      601 accgacaccg tcttccgcgt caccgccacc ggggacggct tcaccctgcg ggaggtgccg
      661 gtggacccgc ccctgaccaa ggtgttcccc gacgacgaat cggacgacga atcggacgac
      721 ggggaggacg gcgacccgga ctcccggacg ttcgtcgcgt acggggacga cggcgacctg
      781 gcgggcttcg tggtcgtctc gtactccggc tggaaccgcc ggctgaccgt cgaggacatc
      841 gaggtcgccc cggagcaccg ggggcacggg gtcgggcgcg cgttgatggg gctcgcgacg
      901 gagttcgccc gcgagcgggg cgccgggcac ctctggctgg aggtcaccaa cgtcaacgca
      961 ccggcgatcc acgcgtaccg gcggatgggg ttcaccctct gcggcctgga caccgccctg
     1021 tacgacggca ccgcctcgga cggcgagcag gcgctctaca tgagcatgcc ctgcccctaa
     1081 tcagtactga caataaaaag attcttgttt tcaagaactt gtcatttgta tagttttttt
     1141 atattgtagt tgttctattt taatcaaatg ttagcgtgat ttatattttt tttcgcctcg
     1201 acatcatctg cccagatgcg aagttaagtg cgcagaaagt aatatcatgc gtcaatcgta
     1261 tgtgaatgct ggtcgctata ctgctgtcga ttcgatacta acgccgccat ccagtgtcga
     1321 aaacgagctc tcgagaaccc ttaatgtcga cagtcgaaca agaagcaggc aaagtttaga
     1381 gcactgcccc tccgcactca aaaaagaaaa aactaggagg aaaataaaat tctcaaccac
     1441 acaaacacat aaacacatac aaatacaaat acaagcttat ttacttgaca tcgcgcgatc
     1501 ttccactatt cagcgccgtc cgccctctct cgtgtttttt gtttacgcga caactatgcg
     1561 aaatccggag caacgggcaa ccgtttgggg aaagaccaca cccacgcgcg atcgccatgg
     1621 caacgaggtc gcacacgccc cacacccaga cctccctgcg agcgggcatg ggtacaatgt
     1681 ccccgttgcc acagacacca cttcgtagca cagcgcagag cgtagcgtgt tgttgctgct
     1741 gacaaaagaa aatttttctt agcaaagcaa aggaggggaa gcacgggcag atagcaccgt
     1801 accataccct tggaaactcg aaatgaacga agcaggaaat gagagaatga gagttttgta
     1861 ggtatatata gcggtagtgt ttgcgcgtta ccatcatctt ctggatctat ctattgttct
     1921 tttcctcatc actttcccct ttttcgctct tcttcttgtc ttttatttct ttcttttttt
     1981 taattgttcc ctcgattggc tatctaccaa agaatccaaa cttaatacac gtatttattt
     2041 gtccaattac catggacaag aaatacagca tcggcctaga tatcgggact aacagcgtcg
     2101 gatgggcggt aattacggac gagtataagg tacctagcaa gaagtttaaa gtactgggaa
     2161 acactgaccg tcactcaatc aagaagaacc taattggggc tttgttattc gactcaggtg
     2221 agacggctga agctacccgt cttaagagga cggcaagaag gcgttatact agacgtaaga
     2281 acagaatttg ttatctacaa gagatctttt caaatgaaat ggcaaaggtt gacgactctt
     2341 tctttcatag actggaggaa tcatttctgg ttgaggagga caaaaagcac gagcgtcacc
     2401 caatcttcgg taatatagtt gatgaagttg cttaccatga gaaatacccg acgatatatc
     2461 atttgagaaa gaaactagta gactcaacag acaaggccga cctgcgttta atctaccttg
     2521 cccttgctca tatgataaaa ttcaggggcc atttcttgat agaaggtgac ttgaaccccg
     2581 acaattctga cgttgacaaa ctatttattc aattggttca gacctacaac cagctgttcg
     2641 aagagaatcc cataaatgca tctggagtgg atgctaaagc aatcttatcc gcaagattat
     2701 caaagtccag aagattggag aatctgatcg ctcagttacc gggcgagaaa aagaatggtt
     2761 tgttcggtaa tttgattgct cttagcttag gtctaacgcc caacttcaag tccaacttcg
     2821 atttggctga ggacgccaag ttgcagttgt ctaaggacac ttatgatgac gatttggata
     2881 atttgttagc ccagataggg gaccaatatg cagacttatt tcttgcggcg aagaacctat
     2941 cagacgcgat cttattgagt gacattttaa gggttaatac tgaaattacg aaggccccac
     3001 tgtcagcgtc aatgattaag cgttatgatg agcatcatca ggacttgact ctactaaagg
     3061 cgttggtcag acagcaactg ccagagaagt ataaagagat cttcttcgat cagtctaaaa
     3121 atgggtatgc gggctatata gatggtggtg cttcacaaga agagttttac aagtttataa
     3181 aacccatcct tgaaaaaatg gacggtacgg aagagctatt agtcaaactt aacagggagg
     3241 acctactgag gaaacagaga acgttcgaca atggatctat accgcaccag atacacttag
     3301 gggagttaca tgctatcctt cgtcgtcaag aagattttta cccgttcctt aaagataaca
     3361 gagaaaaaat tgagaagatt ctgacattca ggataccata ttacgtaggt cccttagcca
     3421 gaggaaattc ccgttttgcg tggatgactc gtaaatcaga agaaaccata actccttgga
     3481 atttcgaaga agtggtggac aaaggcgctt ccgctcaaag tttcatcgaa cgtatgacca
     3541 acttcgataa aaacctgccg aatgagaaag ttttgcctaa gcattctttg ctgtacgagt
     3601 atttcacggt ctataacgaa ctaacaaagg ttaagtacgt aaccgaggga atgaggaagc
     3661 cagcgttttt atctggcgag caaaaaaaag ccatagttga tctattgttc aagacgaaca
     3721 ggaaagtaac agttaaacaa ctaaaagagg actattttaa gaagattgaa tgctttgact
     3781 ctgtcgagat tagcggggtg gaggatcgtt tcaacgcgag cctagggact taccatgatc
     3841 tattaaagat cataaaagat aaagactttc tagataacga agagaatgaa gatatattag
     3901 aagacatagt cctgacttta acattgttcg aagacaggga aatgatcgaa gaaaggctta
     3961 agacatacgc gcatcttttc gacgataaag tcatgaaaca gcttaagcgt aggcgttata
     4021 cggggtgggg gagactgagt aggaagctga tcaatgggat cagggacaag caatcaggta
     4081 aaacaatcct agatttttta aagtccgacg ggttcgcaaa tcgtaatttt atgcagttaa
     4141 tccatgatga cagtttaacg tttaaagagg acatacaaaa ggctcaagtg agtggacagg
     4201 gagatagcct gcacgagcac atcgctaatc tggccggctc tcctgcgatt aaaaagggta
     4261 tactgcaaac cgtaaaagtg gtagacgaat tggttaaggt catgggcagg cataagccgg
     4321 agaacatagt aatcgagatg gcgagagaaa atcagacgac tcaaaagggt cagaagaact
     4381 ctagagagcg tatgaagcgt attgaagagg gaatcaagga gcttggctcc caaatcctaa
     4441 aagagcatcc cgtggagaat acccaattgc aaaatgaaaa actgtatttg tattacttgc
     4501 aaaacggcag ggacatgtac gttgatcaag aattggatat caatagattg tcagactatg
     4561 atgttgacca tattgttccc cagtctttcc taaaagatga ctctatagat aacaaggtct
     4621 tgactagaag cgataaaaac agggggaagt ctgacaacgt cccctctgag gaagtcgtga
     4681 agaagatgaa gaactactgg cgtcagctat tgaatgcgaa attaattacg caacgtaaat
     4741 tcgataacct aaccaaggca gagcgtgggg ggctttcaga gctagataaa gcaggtttta
     4801 taaagaggca actagtagag actcgtcaaa ttaccaagca tgttgcccag atattagact
     4861 cccgtatgaa caccaaatat gacgagaacg acaagctgat cagggaggta aaggttatca
     4921 cgttaaagtc caagttagtt tctgacttca ggaaggattt tcagttctac aaggtacgtg
     4981 aaattaataa ttatcaccac gcgcacgacg cttatttgaa tgctgttgtg ggaactgcat
     5041 taattaaaaa gtacccaaaa cttgagtctg agtttgtgta tggcgactat aaggtttatg
     5101 atgtcaggaa aatgattgcc aagtctgagc aggagatagg aaaggctacc gcaaagtatt
     5161 ttttttactc caacattatg aatttcttta aaacggaaat tactctggcg aatggtgaaa
     5221 ttaggaaaag acctttaata gagaccaatg gtgagacagg cgagattgtc tgggataagg
     5281 gccgtgattt cgcaaccgtt agaaaagtac tatccatgcc acaagtgaac atagttaaaa
     5341 aaacggaagt ccaaacagga ggctttagta aggagagcat tcttcccaaa aggaacagtg
     5401 ataaattaat cgcgaggaaa aaagactggg atccgaagaa gtatggggga ttcgactctc
     5461 caaccgtggc ctattccgta ttggtcgtcg caaaagtaga aaaaggtaag tccaagaagc
     5521 tgaaaagcgt caaagagtta ttgggcatca caatcatgga gaggtcatcc ttcgagaaaa
     5581 accctatcga tttcttagaa gcaaaaggat ataaggaggt taaaaaagac ctaattataa
     5641 aactacctaa gtattcctta ttcgagttgg agaacggcag aaagaggatg cttgccagcg
     5701 caggggagtt acagaaaggg aacgagctag ccctgccctc aaaatatgtg aatttcttat
     5761 acttagcctc acactacgaa aagttaaaag ggtctccgga agataatgag caaaaacagc
     5821 tattcgttga acagcacaaa cattatttag acgagatcat tgaacaaatt tcagagtttt
     5881 caaaaagggt catactagct gatgctaact tagacaaagt tctttccgct tataacaaac
     5941 atagggataa gccgatcaga gagcaagctg aaaacataat tcatttattt acgcttacga
     6001 acctgggagc cccggcggcc ttcaagtact tcgataccac aatcgaccgt aagaggtaca
     6061 catccactaa ggaagtgtta gatgcgacac taattcatca gagtattact ggcctgtatg
     6121 aaacgcgtat agatttgagc caattaggcg gcgattcaag agccgatccg aagaaaaaaa
     6181 ggaaagtata aatccgctct aaccgaaaag gaaggagtta gacaacctga agtctaggtc
     6241 cctatttatt tttttatagt tatgttagta ttaagaacgt tatttatatt tcaaattttt
     6301 cttttttttc tgtacagacg cgtgtacgca tgtaacatta tactgaaaac cttgcttgag
     6361 aaggttttgg gacgctcgaa gccgcggtga gcaaaaggcc agcaaaaggc caggaaccgt
     6421 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     6481 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     6541 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     6601 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     6661 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     6721 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     6781 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     6841 acagagttct tgaagtggtg gcctaactac ggctacacta gaaggacagt atttggtatc
     6901 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     6961 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     7021 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     7081 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     7141 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     7201 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     7261 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     7321 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     7381 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     7441 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     7501 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     7561 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     7621 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     7681 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     7741 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     7801 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     7861 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     7921 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     7981 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     8041 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     8101 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     8161 gttccgcgca catttccccg aaaagtgcca cctgaacgaa gcatctgtgc ttcattttgt
     8221 agaacaaaaa tgcaacgcga gagcgctaat ttttcaaaca aagaatctga gctgcatttt
     8281 tacagaacag aaatgcaacg cgaaagcgct attttaccaa cgaagaatct gtgcttcatt
     8341 tttgtaaaac aaaaatgcaa cgcgagagcg ctaatttttc aaacaaagaa tctgagctgc
     8401 atttttacag aacagaaatg caacgcgaga gcgctatttt accaacaaag aatctatact
     8461 tcttttttgt tctacaaaaa tgcatcccga gagcgctatt tttctaacaa agcatcttag
     8521 attacttttt ttctcctttg tgcgctctat aatgcagtct cttgataact ttttgcactg
     8581 taggtccgtt aaggttagaa gaaggctact ttggtgtcta ttttctcttc cataaaaaaa
     8641 gcctgactcc acttcccgcg tttactgatt actagcgaag ctgcgggtgc attttttcaa
     8701 gataaaggca tccccgatta tattctatac cgatgtggat tgcgcatact ttgtgaacag
     8761 aaagtgatag cgttgatgat tcttcattgg tcagaaaatt atgaacggtt tcttctattt
     8821 tgtctctata tactacgtat aggaaatgtt tacattttcg tattgttttc gattcactct
     8881 atgaatagtt cttactacaa tttttttgtc taaagagtaa tactagagat aaacataaaa
     8941 aatgtagagg tcgagtttag atgcaagttc aaggagcgaa aggtggatgg gtaggttata
     9001 tagggatata gcacagagat atatagcaaa gagatacttt tgagcaatgt ttgtggaagc
     9061 ggtattcgca atattttagt agctcgttac agtccggtgc gtttttggtt ttttgaaagt
     9121 gcgtcttcag agcgcttttg gttttcaaaa gcgctctgaa gttcctatac tttctagaga
     9181 ataggaactt cggaatagga acttcaaagc gtttccgaaa acgagcgctt ccgaaaatgc
     9241 aacgcgagct gcgcacatac agctcactgt tcacgtcgca cctatatctg cgtgttgcct
     9301 gtatatatat atacatgaga agaacggcat agtgcgtgtt tatgcttaaa tgc
//
