V"V"AID EU855180; SV 1; circular; other DNA; STD; SYN; 10595 BP. AC EU855180; DT 03-AUG-2008 (Rel. 96, Created) DT 03-AUG-2008 (Rel. 96, Last updated, Version 1) DE Cloning vector pAW8_GST, complete sequence. KW . OS Cloning vector pAW8_GST OC other sequences; artificial sequences; vectors. RN [1] RP 1-10595 RX DOI; 10.1016/j.gene.2007.09.024. RX PUBMED; 18054176. RA Watson A.T., Garcia V., Bone N., Carr A.M., Armstrong J.; RT "Gene tagging and gene replacement using recombinase-mediated cassette RT exchange in Schizosaccharomyces pombe"; RL Gene 407(1-2):63-74(2008). RN [2] RP 1-10595 RA Watson A.T.; RT ; RL Submitted (27-JUN-2008) to the EMBL/GenBank/DDBJ databases. RL Genome Damage and Stability Centre, University of Sussex, Falmer, Brighton, RL East Sussex BN1 9RQ, UK FH Key Location/Qualifiers FT source 1..10595 FT /organism="Cloning vector pAW8_GST" FT /mol_type="other DNA" FT /db_xref="taxon:550442" FT misc_feature 1..3873 FT /note="Cre-expression plasmid pAW8 backbone" FT misc_recomb 3874..3907 FT /note="loxP" FT gene 3916..4590 FT /gene="gst" FT CDS 3916..4590 FT /codon_start=1 FT /transl_table=11 FT /gene="gst" FT /product="glutathione-S-transferase affinity tag" FT /note="GST affinity tag" FT /protein_id="ACF75943.1" FT /translation="MSPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNKK FT FELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERAEISMLEGAVLDIRY FT GVSRIAYSKDFETLKVDFLSKLPEMLKMFEDRLCHKTYLNGDHVTHPDFMLYDALDVVL FT YMDPMCLDAFPKLVCFKKRIEAIPQIDKYLKSSKYIAWPLQGWQATFGGGDHPPKSDLV FT PR" FT terminator 4605..4833 FT /note="Saccharomyces cerevisiae ADH1" FT gene 5223..6032 FT /gene="kanR" FT CDS 5223..6032 FT /codon_start=1 FT /transl_table=11 FT /gene="kanR" FT /product="kanamycin and geneticin G418 resistance protein" FT /protein_id="ACF75944.1" FT /translation="MGKEKTHVSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYGKP FT DAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGKTA FT FQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDASD FT FDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGIAD FT RYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF" FT misc_recomb 6280..6313 FT /note="loxM3" FT misc_feature 6314..10595 FT /note="Cre-expression plasmid pAW8 backbone" SQ Sequence 10595 BP; 2987 A; 2243 C; 2220 G; 3145 T; 0 other; caaaaagtta ggtgtaacag aaaatcatga aactatgatt tctaatttat atattggagg 60 attttctcta aaaaaaaaaa tacaacaaat aaaaaacact caatgacctg accatttgat 120 ggagtttaag tcaatacctt cttgaaccat ttcccataat ggtgaaagtt ccctcaagaa 180 ttttactctg tcagaaacgg ccttaacgac gtagtcgatc cggacgatgg ccaaccttcc 240 aattcattaa atcgagtcct ccatcagatg atttacaagg ttttgcatag aatccctaag 300 taagagaagt tgttagtgat ttcttcaagt ttactaaaat ttacatacaa atggatgatc 360 tcttctccat tgtttccttt cttcttgcaa acgcgtttta caaagagatg acatgctatt 420 ttctgattat ttttttctat agttttctat tttgtacgtc cttgggagtc cgaaatgtaa 480 aatcggggta tggttagtag cggcagcgtt cattatggta tataaaagat aaaaagtagg 540 ggcaagcgag acagaatggg atacaagggc atcgtctata attatagcta aaaattgtat 600 tttaatttgt attttttgta attttatttc tcattgtttt acttaaaatg aatgcgaatt 660 agagaaaact tcaacgaaat gtcaaaataa gctcagcaaa atatacaatt ttagggaaag 720 cgatcagcaa cttattctcc ggtgtagatt ctttttcttc ataaattcca acaatgtaac 780 tgccaattga tttttctcta gatgctaata aattgaaact aatgaatata ataacacagg 840 ttttgataaa ttgtaaactc ttactcaaaa aaagaatgtt aaaactaatt taaccttttt 900 ataaaggctc atggcgttaa agaggatagt aaaaattcta caatttttta atttaatgtc 960 acaattgtct tgcgaatata attctattaa ttttcgctaa ctgctgtttt agtactatta 1020 ctttcaatat gccttatatg caactttaat tcatgaagaa tgcaatattt ttgcaatctt 1080 tcttgtactg agcgcttttt atcatttagt actcattatt taatttttca gtaaagggca 1140 ataaataaat ttgtagaaga tgcaatgtaa tctcctctat ccttttgctc atatgtttat 1200 gagtatacct agtctagaaa ggcttgtatt taaaatatta ttcaataaaa attcacaatt 1260 tttacgacat gtgctaccat tcacttaact tcctgattat aaaattggtt cgtttatact 1320 aattacttaa gtacctttaa ctaaacaaaa tgcctatata tatattaatt tacaatgagt 1380 gtcagataag tcactatgtc cgagtggtta aggagttaga ctcgccatcg atggtatggt 1440 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa 1500 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg 1560 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga 1620 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt 1680 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt 1740 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat 1800 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt 1860 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg 1920 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga 1980 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc 2040 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac 2100 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg 2160 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca 2220 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg 2280 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg 2340 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg 2400 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag 2460 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg 2520 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct 2580 cccgtatcgt agttatctac acgacgggga gtcaggccac tatggatgaa cgaaatagac 2640 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact 2700 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga 2760 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt 2820 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct 2880 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc 2940 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc 3000 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc 3060 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg 3120 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt 3180 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg 3240 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg 3300 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt 3360 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag 3420 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt 3480 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta 3540 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt 3600 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc 3660 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca 3720 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc 3780 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg 3840 accatgatta cgccaagctt gcatggcggc cgcataactt cgtatagcat acattatacg 3900 aagttatgca tgctcatgtc ccctatacta ggttattgga aaattaaggg ccttgtgcaa 3960 cccactcgac ttcttttgga atatcttgaa gaaaaatatg aagagcattt gtatgagcgc 4020 gatgaaggtg ataaatggcg aaacaaaaag tttgaattgg gtttggagtt tcccaatctt 4080 ccttattata ttgatggtga tgttaaatta acacagtcta tggccatcat acgttatata 4140 gctgacaagc acaacatgtt gggtggttgt ccaaaagagc gtgcagagat ttcaatgctt 4200 gaaggagcgg ttttggatat tagatacggt gtttcgagaa ttgcatatag taaagacttt 4260 gaaactctca aagttgattt tcttagcaag ctacctgaaa tgctgaaaat gttcgaagat 4320 cgtttatgtc ataaaacata tttaaatggt gatcatgtaa cccatcctga cttcatgttg 4380 tatgacgctc ttgatgttgt tttatacatg gacccaatgt gcctggatgc gttcccaaaa 4440 ttagtttgtt ttaaaaaacg tattgaagct atcccacaaa ttgataagta cttgaaatcc 4500 agcaagtata tagcatggcc tttgcagggc tggcaagcca cgtttggtgg tggcgaccat 4560 cctccaaaat cggatctggt tccgcgttga ggcgcgccag atctacttct aaataagcga 4620 atttcttatg atttatgatt tttattatta aataagttat aaaaaaaata agtgtataca 4680 aattttaaag tgactcttag gttttaaaac gaaaattctt attcttgagt aactctttcc 4740 tgtaggtcag gttgctttct caggtatagt atgaggtcgc tcttattgac cacacctcta 4800 ccggcagatc cgctagggat aacagggtaa tatagatctg tttagcttgc ctcgtccccg 4860 ccgggtcacc cggccagcga catggaggcc cagaataccc tccttgacag tcttgacgtg 4920 cgcagctcag gggcatgatg tgactgtcgc ccgtacattt agcccataca tccccatgta 4980 taatcatttg catccataca ttttgatggc cgcacggcgc gaagcaaaaa ttacggctcc 5040 tcgctgcaga cctgcgagca gggaaacgct cccctcacag acgcgttgaa ttgtccccac 5100 gccgcgcccc tgtagagaaa tataaaaggt taggatttgc cactgaggtt cttctttcat 5160 atacttcctt ttaaaatctt gctaggatac agttctcaca tcacatccga acataaacaa 5220 ccatgggtaa ggaaaagact cacgtttcga ggccgcgatt aaattccaac atggatgctg 5280 atttatatgg gtataaatgg gctcgcgata atgtcgggca atcaggtgcg acaatctatc 5340 gattgtatgg gaagcccgat gcgccagagt tgtttctgaa acatggcaaa ggtagcgttg 5400 ccaatgatgt tacagatgag atggtcagac taaactggct gacggaattt atgcctcttc 5460 cgaccatcaa gcattttatc cgtactcctg atgatgcatg gttactcacc actgcgatcc 5520 ccggcaaaac agcattccag gtattagaag aatatcctga ttcaggtgaa aatattgttg 5580 atgcgctggc agtgttcctg cgccggttgc attcgattcc tgtttgtaat tgtcctttta 5640 acagcgatcg cgtatttcgt ctcgctcagg cgcaatcacg aatgaataac ggtttggttg 5700 atgcgagtga ttttgatgac gagcgtaatg gctggcctgt tgaacaagtc tggaaagaaa 5760 tgcataagct tttgccattc tcaccggatt cagtcgtcac tcatggtgat ttctcacttg 5820 ataaccttat ttttgacgag gggaaattaa taggttgtat tgatgttgga cgagtcggaa 5880 tcgcagaccg ataccaggat cttgccatcc tatggaactg cctcggtgag ttttctcctt 5940 cattacagaa acggcttttt caaaaatatg gtattgataa tcctgatatg aataaattgc 6000 agtttcattt gatgctcgat gagtttttct aatcagtact gacaataaaa agattcttgt 6060 tttcaagaac ttgtcatttg tatagttttt ttatattgta gttgttctat tttaatcaaa 6120 tgttagcgtg atttatattt tttttcgcct cgacatcatc tgcccagatg cgaagttaag 6180 tgcgcagaaa gtaatatcat gcgtcaatcg tatgtgaatg ctggtcgcta tactgctgtc 6240 gattcgatac taacgccgcc atccagtcga cgaactagta taacttcgta tatggtatta 6300 tatacgaagt tatgcggccg ctgaattcac tggccgtcgt tttacaacgt cgtgactggg 6360 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc 6420 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg 6480 aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac 6540 catcgtccgg atcaatgaag tagatgagtt tactacctgt atatactttt ttttttgtca 6600 aagtcacttt tatggcattt ccatgttgaa ggattagcca cagcactaaa ttttcagcgt 6660 atgattgctt ttaaatattt aattttcatc gttttttaat taatattcaa aacgatttaa 6720 tgccttctac atctgaagtc aagctcatag actcgggttg ggtagctaaa actacatcta 6780 acacgaaggg gtatagcctt cttaaagtat actcatgtat aaacgttttg ggttcatatt 6840 tttgggccac gtggtttata aaaattctta actacaccac tcggtatccc gcacccgtct 6900 acgtttctac gatttcgagc taatattgag taaagtgatt agcaaaaaaa ataaaagtac 6960 tcgttgtcgg agatcaagaa tttttctatt atctcatcta aaccactttc taaaagcgaa 7020 aaacaaaatc gtaatatgca gcttgaatgg gcttccatag tttgaaagaa aaaccctagc 7080 agtactggca agggagacat tccttttacc tggctaatcg ccatcttcca gcaggcgcac 7140 cattgcccct gtttcactat ccaggttacg gatatagttc atgacaatat ttacattggt 7200 ccagccacca gcttgcatga tctccggtat tgaaactcca gcgcgggcca tatctcgcgc 7260 ggctccgaca cgggcactgt gtccagacca ggccaggtat ctctgaccag agtcatcctt 7320 agcgccgtaa atcaatcgat gagttgcttc aaaaatccct tccagggcgc gagttgatag 7380 ctggctggtg gcagatggcg cggcaacacc attttttctg acccggcaaa acaggtagtt 7440 attcggatca tcagctacac cagagacgga aatccatcgc tcgaccagtt tagttacccc 7500 caggctaagt gccttctcta cacctgcggt gctaaccagc gttttcgttc tgccaatatg 7560 gattaacatt ctcccaccgt cagtacgtga gatatcttta accctgatcc tggcaatttc 7620 ggctatacgt aacagggtgt tataagcaat ccccagaaat gccagattac gtatatcctg 7680 gcagcgatcg ctattttcca tgagtgaacg aacctggtcg aaatcagtgc gttcgaacgc 7740 tagagcctgt tttgcacgtt caccggcatc aacgttttct tttcggatcc gccgcataac 7800 cagtgaaaca gcattgctgt cacttggtcg tggcagcccg gaccgacgat gaagcatgtt 7860 tagctggccc aaatgttgct ggatagtttt tactgccaga ccgcgcgcct gaagatatag 7920 aagataatcg cgaacatctt caggttctgc gggaaaccat ttccggttat tcaacttgca 7980 ccatgccgcc cacgaccggc aaacggacag aagcattttc caggtatgct cagaaaacgc 8040 ctggcgatcc ctgaacatgt ccatcaggtt cttgcgaacc tcatcactcg ttgcatcgac 8100 cggtaacttt tagattaaca tttgtttaaa tatgaaatta ggcaatgtaa catactgcag 8160 gcaaattttg gtgtacggta agtaaattgg acatatgatt taacaaagcg actataagtc 8220 agaaagtgag aatgagattg aaataattaa ttcaacttat tcaattgatg atatgccagg 8280 attcctcttc ctttatatat cttcaatgtt tcgattaaca ggtccattat ccggttttta 8340 gtttcgctgc acattgccga atgacaatcg gattccttta agctttgctt ttgccctgtt 8400 ttctctcccc agtaatcgtt gggtgactga accatccagt agccgagagt tacgtttttc 8460 ctgatttaat aaatatccat atctatttgt ctcgtggtgt tttacagcaa ctaaccgaaa 8520 ttactatttt gcatcatcca accataaaga gacaacaata attctagaaa acatataaca 8580 agtgtttttt tctatttacg tctcatacgc aaagtccact ttaccatgtt ttggaaactt 8640 gtacccaacc attcattttt cgattttgta atgaggctct tttcttctaa caatcattga 8700 ggtcattatc atgtgtcctc acctattgtt taaattgagg cttgcgctat tcccaaaatt 8760 gtgggtatta ttttcatata tcagtatctg caatttcggt tcaactccgg aagctcggta 8820 cccaactgtg ggaatactca ggtatcgtaa gatgcaagag ttcgaatctc ttagcaacca 8880 ttattttttt tctctcaaca taacgagaac acacaggggc gctatcgcac agaatcaaat 8940 tcgatgactg gaaatttttt gttaatttca gaggtcgcct gacgcatata cctttttcaa 9000 ctgaaaaatt gggagaaaaa ggaaaggtga gagcgccgga accggctttt catatagaat 9060 agagaagcgt tcatgactaa atgcttgcat cacaatactt gaagttgaca atattattta 9120 aggacctatt gttttttcca ataggtggtt agcaatcgtc ttactttcta acttttctta 9180 ccttttacat ttcagcaata tatatatata tatatatata tatttcaagg atataccatt 9240 gtaatgtctg cccctaagaa gatcgtcgtt ttgccaggtg accacgttgg tcaagaaatc 9300 acagccgaag ccattaaggt tcttaaagct atttctgatg ttcgttccaa tgtcaagttc 9360 gatttcgaaa atcatttaat tggtggtgct gctatcgatg ctacaggtgt tccacttcca 9420 gatgaggcgc tggaagcctc caagaaggct gatgccgttt tgttaggtgc tgtgggtggt 9480 cctaaatggg gtaccggtag tgttagacct gaacaaggtt tactaaaaat ccgtaaagaa 9540 cttcaattgt acgccaactt aagaccatgt aactttgcat ccgactctct tttagactta 9600 tctccaatca agccacaatt tgctaaaggt actgacttcg ttgttgttag agaattagtg 9660 ggaggtattt actttggtaa gagaaaggaa gacgatggtg atggtgtcgc ttgggatagt 9720 gaacaataca ccgttccaga agtgcaaaga atcacaagaa tggccgcttt catggcccta 9780 caacatgagc caccattgcc tatttggtcc ttggataaag ctaatgtttt ggcctcttca 9840 agattatgga gaaaaactgt ggaggaaacc atcaagaacg aatttcctac attgaaagtt 9900 caacatcaat tgattgattc tgccgccatg atcctagtta agaacccaac ccacctaaat 9960 ggtattataa tcaccagcaa catgtttggt gatatcatct ccgatgaagc ctccgttatc 10020 ccaggctcct tgggtttgtt gccatctgcg tccttggcct ctttgccaga caagaacacc 10080 gcatttggtt tgtacgaacc atgccatggt tccgctccag atttgccaaa gaataaggtc 10140 aaccctatcg ccactatctt gtctgctgca atgatgttga aattgtcatt gaacttgcct 10200 gaagaaggta aagccattga agatgcagtt aaaaaggttt tggatgcagg tatcagaact 10260 ggtgatttag gtggttccaa cagtaccacc gaagtcggtg atgctgtcgc cgaagaagtt 10320 aagaaaatcc ttgcttaaaa agattctctt tttttgtgat atttgtacat aaactttata 10380 aatgaaattc ataatagaaa cgacacgaaa ttacaaaatg gaatatgttc atagggtaga 10440 cgaaactata tacgcaatct acatacattt atcaagaagg agaaaaagga ggatgtaaag 10500 gaatacaggt aagcaaattg atactaatgg ctcaacgtga taaggaaaaa gaattgcact 10560 ttaacattaa tattgacaag gaggagggca tcaca 10595 c)pAW8_GST"Times New Roman V"MluI(392),XbaI(798),ScaI(1014) Arialc)of] /] V"+ScaI(1111),NdeI(1192),XbaI(1214),ClaI(1429) Arialc)?FL|LL| V"ScaI ArialPc) V"EPvuII(3677),HindIII(3856),NotI(3868),EagI(3868),SwaI(4344),BclI(4351) Arialc)Yahh' V"CScaI(4490),AscI(4593),BglII(4600),BglII(4835),PstI(5049),MluI(5082) Arialc)  ##Mk V".NcoI(5222),NruI(5306),ClaI(5340),HindIII(5767) Arialc)"5EEu V"YScaI(6038),SalI(6267),SpeI(6275),NotI(6316),EagI(6316),EcoRI(6324),PvuII(6416),PacI(6701) Arialc)+%n V",ScaI(6959),ScaI(7084),ClaI(7337),EcoRV(7594) Arialc) V"FSexAI(7713),BamHI(7786),NruI(7931),PstI(8160),NdeI(8194),HindIII(8381) Arialc)@@@rd V"XbaI(8564),KpnI(8822) Arialt!c) V"!ClaI(9396),KpnI(9495),EcoRV(9994) Arial%c)~vRv V"NcoI Arialy'c)h`eVbLF@~LV"Cc)"8 pGEX_3_primer"Times New RomanV"CɆc) AmpR_promoter"Times New RomanV"Cc)  Ampicillin"Times New RomanV"Cc)y ORF 3"Times New RomanV"c)   pBR322_ori"Times New RomanV"c) lac_promoter"Times New RomanV"Cc)M13_pUC_rev_primer"Times New RomanV"CɆc)M13_reverse_primer"Times New RomanV"CɆc)ORF 1"Times New RomanV"Cc)"CloxP"Times New RomanV"Cc)Lgst"Times New RomanV"Cc)@XpHybLex_rev_primer"Times New RomanV"c)gkanR"Times New RomanV"CɆc)M13_forward20_primer"Times New RomanV"c)M13_pUC_fwd_primer"Times New RomanV"Cc)ORF 1"Times New RomanV"Cc)$b(ORF 2"Times New Roman V"test"Times New Romanc)q^GG V"test"Times New RomanSc)<2"Iq" V"test"Times New Romanc)$*88