LOCUS Exported 9128 bp ds-DNA circular SYN 13-MAY-2021 DEFINITION Cre-expression plasmid. ACCESSION . VERSION . KEYWORDS pAW5 SOURCE synthetic DNA construct ORGANISM synthetic DNA construct REFERENCE 1 (bases 1 to 9128) AUTHORS Watson AT1, Garcia V, Bone N, Carr AM, Armstrong J. TITLE Gene tagging and gene replacement using recombinase-mediated cassette exchange in Schizosaccharomyces pombe. JOURNAL Gene. 2008 Jan 15;407(1-2):63-74 PUBMED 18054176 REFERENCE 2 (bases 1 to 9128) AUTHORS . TITLE Direct Submission JOURNAL Exported May 13, 2021 from SnapGene Server 1.1.58 http://www.snapgene.com FEATURES Location/Qualifiers source 1..9128 /organism="synthetic DNA construct" /mol_type="other DNA" rep_origin 575..1356 /label=ars1 /note="Schizosaccharomyces pombe autonomously replicating sequence ars1" primer_bind complement(1451..1470) /label=pRS-marker /note="pRS vectors, use to sequence yeast selectable marker" primer_bind 1570..1592 /label=pGEX 3' /note="pGEX vectors, reverse primer" primer_bind complement(1630..1648) /label=pBRforEco /note="pBR322 vectors, upsteam of EcoRI site, forward primer" promoter 1716..1820 /gene="bla" /label=AmpR promoter CDS 1821..2681 /codon_start=1 /gene="bla" /product="beta-lactamase" /label=AmpR /note="confers resistance to ampicillin, carbenicillin, and related antibiotics" /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS LIKHW" primer_bind complement(2039..2058) /label=Amp-R /note="Ampicillin resistance gene, reverse primer" rep_origin 2852..3440 /direction=RIGHT /label=ori /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of replication" primer_bind 3341..3360 /label=pBR322ori-F /note="pBR322 origin, forward primer" primer_bind 3594..3611 /label=L4440 /note="L4440 vector, forward primer" protein_bind 3728..3749 /label=CAP binding site /bound_moiety="E. coli catabolite activator protein" /note="CAP binding activates transcription in the presence of cAMP." promoter 3764..3794 /label=lac promoter /note="promoter for the E. coli lac operon" protein_bind 3802..3818 /label=lac operator /bound_moiety="lac repressor encoded by lacI" /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)." primer_bind 3807..3829 /label=M13/pUC Reverse /note="In lacZ gene" primer_bind 3826..3842 /label=M13 rev /note="common sequencing primer, one of multiple similar variants" primer_bind 3826..3842 /label=M13 Reverse /note="In lacZ gene. Also called M13-rev" CDS 3838..4149 /codon_start=1 /gene="lacZ fragment" /product="LacZ-alpha fragment of beta-galactosidase" /label=lacZ-alpha /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIPSSGSSHY" misc_feature 3855..3911 /label=MCS /note="pUC18/19 multiple cloning site" primer_bind complement(3912..3929) /label=M13 Forward /note="In lacZ gene. Also called M13-F20 or M13 (-21) Forward" primer_bind complement(3912..3928) /label=M13 fwd /note="common sequencing primer, one of multiple similar variants" primer_bind complement(3921..3943) /label=M13/pUC Forward /note="In lacZ gene" CDS complement(5137..6168) /codon_start=1 /product="site-specific recombinase" /label=Cre /note="Cre is a site-specific recombinase from bacteriophage P1. Recombination occurs at loxP sequences." /translation="MSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE DGD" CDS 7777..8871 /codon_start=1 /gene="S. cerevisiae LEU2" /product="3-isopropylmalate dehydrogenase, required for leucine biosynthesis" /label=LEU2 /note="yeast auxotrophic marker" /translation="MSAPKKIVVLPGDHVGQEITAEAIKVLKAISDVRSNVKFDFENHL IGGAAIDATGVPLPDEALEASKKADAVLLGAVGGPKWGTGSVRPEQGLLKIRKELQLYA NLRPCNFASDSLLDLSPIKPQFAKGTDFVVVRELVGGIYFGKRKEDDGDGVAWDSEQYT VPEVQRITRMAAFMALQHEPPLPIWSLDKANVLASSRLWRKTVEETIKNEFPTLKVQHQ LIDSAAMILVKNPTHLNGIIITSNMFGDIISDEASVIPGSLGLLPSASLASLPDKNTAF GLYEPCHGSAPDLPKNKVNPIATILSAAMMLKLSLNLPEEGKAIEDAVKKVLDAGIRTG DLGGSNSTTEVGDAVAEEVKKILA" ORIGIN 1 caaaaagtta ggtgtaacag aaaatcatga aactatgatt tctaatttat atattggagg 61 attttctcta aaaaaaaaaa tacaacaaat aaaaaacact caatgacctg accatttgat 121 ggagtttaag tcaatacctt cttgaaccat ttcccataat ggtgaaagtt ccctcaagaa 181 ttttactctg tcagaaacgg ccttaacgac gtagtcgatc cggacgatgg ccaaccttcc 241 aattcattaa atcgagtcct ccatcagatg atttacaagg ttttgcatag aatccctaag 301 taagagaagt tgttagtgat ttcttcaagt ttactaaaat ttacatacaa atggatgatc 361 tcttctccat tgtttccttt cttcttgcaa acgcgtttta caaagagatg acatgctatt 421 ttctgattat ttttttctat agttttctat tttgtacgtc cttgggagtc cgaaatgtaa 481 aatcggggta tggttagtag cggcagcgtt cattatggta tataaaagat aaaaagtagg 541 ggcaagcgag acagaatggg atacaagggc atcgtctata attatagcta aaaattgtat 601 tttaatttgt attttttgta attttatttc tcattgtttt acttaaaatg aatgcgaatt 661 agagaaaact tcaacgaaat gtcaaaataa gctcagcaaa atatacaatt ttagggaaag 721 cgatcagcaa cttattctcc gttgtagatt ctttttcttc ataaattcca acaatgtaac 781 tgccaattga tttttctcta gatgctaata aattgaaact aatgaatata ataacacagg 841 ttttgataaa ttgtaaactc ttactcaaaa aaagaatgtt aaaactaatt taaccttttt 901 ataaaggctc atggcgttaa agaggatagt aaaaattcta caatttttta atttaatgtc 961 acaattgtct tgcgaatata attctattaa ttttcgctaa ctgctgtttt agtactatta 1021 ctttcaatat gccttatatg caactttaat tcatgaagaa tgcaatattt ttgcaatctt 1081 tcttgtactg agcgcttttt atcatttagt actcattatt taatttttca gtaaagggca 1141 ataaataaat ttgtagaaga tgcaatgtaa tctcctctat ccttttgctc atatgtttat 1201 gagtatacct agtctagaaa ggcttgtatt taaaatatta ttcaataaaa attcacaatt 1261 tttacgacat gtgctaccat tcacttaact tcctgattat aaaattggtt cgtttatact 1321 aattacttaa gtacctttaa ctaaacaaaa tgcctatata tatattaatt tacaatgagt 1381 gtcagataag tcactatgtc cgagtggtta aggagttaga ctcgccatcg atggtatggt 1441 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa 1501 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg 1561 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga 1621 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt 1681 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt 1741 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat 1801 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt 1861 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg 1921 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga 1981 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc 2041 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac 2101 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg 2161 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca 2221 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg 2281 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg 2341 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg 2401 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag 2461 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg 2521 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct 2581 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac 2641 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact 2701 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga 2761 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt 2821 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct 2881 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc 2941 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc 3001 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc 3061 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg 3121 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt 3181 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg 3241 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg 3301 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt 3361 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag 3421 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt 3481 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta 3541 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt 3601 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc 3661 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca 3721 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc 3781 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg 3841 accatgatta cgccaagctt gcatgcctgc aggtcgactc tagaggatcc ccgggtaccg 3901 agctcgaatt cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc 3961 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc 4021 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggcg cctgatgcgg 4081 tattttctcc ttacgcatct gtgcggtatt tcacaccgca taccatcgtc cggaagctcg 4141 cattactaat agaaaggatt atttcacttc taattacaca aattccgtat aactaacaag 4201 tagtgaaagt ccaagaagta gtctcagagc aaaagccaga ttgaatagaa ttaagaaggc 4261 gctaacaggt ttgggtgata aaacatttat agaatatctt atacatagta ctaaaattta 4321 aatatccagc aagatgtatc catgaaaaca atcacaaagc aaaagacgac gagcgaactt 4381 ttacgatctc aatccgacat ttttgcctcg taaagttttt tttttcagtt ctaatttgga 4441 ctaattctcc caaagcaaaa tgcctctttt tcatccatgc ggccaatctt gtcgaatttc 4501 aatagtatta cttattgaat gttaaactac gatttagcaa taaatattcc tccttccaca 4561 tgctgtaaac aaagtcaatg aagtagatga gtttactacc tgtatatact tttttttttg 4621 tcaaagtcac ttttatggca tttccatgtt gaaggattag ccacagcact aaattttcag 4681 cgtatgattg cttttaaata tttaattttc atcgtttttt aattaatatt caaaacgatt 4741 taatgccttc tacatctgaa gtcaagctca tagactcggg ttgggtagct aaaactacat 4801 ctaacacgaa ggggtatagc cttcttaaag tatactcatg tataaacgtt ttgggttcat 4861 atttttgggc cacgtggttt ataaaaattc ttaactacac cactcggtat cccgcacccg 4921 tctacgtttc tacgatttcg agctaatatt gagtaaagtg attagcaaaa aaaataaaag 4981 tactcgttgt cggagatcaa gaatttttct attatctcat ctaaaccact ttctaaaagc 5041 gaaaaacaaa atcgtaatat gcagcttgaa tgggcttcca tagtttgaaa gaaaaaccct 5101 agcagtactg gcaagggaga cattcctttt acctggctaa tcgccatctt ccagcaggcg 5161 caccattgcc cctgtttcac tatccaggtt acggatatag ttcatgacaa tatttacatt 5221 ggtccagcca ccagcttgca tgatctccgg tattgaaact ccagcgcggg ccatatctcg 5281 cgcggctccg acacgggcac tgtgtccaga ccaggccagg tatctctgac cagagtcatc 5341 cttagcgccg taaatcaatc gatgagttgc ttcaaaaatc ccttccaggg cgcgagttga 5401 tagctggctg gtggcagatg gcgcggcaac accatttttt ctgacccggc aaaacaggta 5461 gttattcgga tcatcagcta caccagagac ggaaatccat cgctcgacca gtttagttac 5521 ccccaggcta agtgccttct ctacacctgc ggtgctaacc agcgttttcg ttctgccaat 5581 atggattaac attctcccac cgtcagtacg tgagatatct ttaaccctga tcctggcaat 5641 ttcggctata cgtaacaggg tgttataagc aatccccaga aatgccagat tacgtatatc 5701 ctggcagcga tcgctatttt ccatgagtga acgaacctgg tcgaaatcag tgcgttcgaa 5761 cgctagagcc tgttttgcac gttcaccggc atcaacgttt tcttttcgga tccgccgcat 5821 aaccagtgaa acagcattgc tgtcacttgg tcgtggcagc ccggaccgac gatgaagcat 5881 gtttagctgg cccaaatgtt gctggatagt ttttactgcc agaccgcgcg cctgaagata 5941 tagaagataa tcgcgaacat cttcaggttc tgcgggaaac catttccggt tattcaactt 6001 gcaccatgcc gcccacgacc ggcaaacgga cagaagcatt ttccaggtat gctcagaaaa 6061 cgcctggcga tccctgaaca tgtccatcag gttcttgcga acctcatcac tcgttgcatc 6121 gaccggtaat gcaggcaaat tttggtgtac ggtaagtaaa ttggacatat gatttaacaa 6181 agcgactata agtcagaaag tgagaatgag attgaaataa ttaattcaac ttattcaatt 6241 gatgatatgc caggattcct cttcctttat cttcaatgtt tcgattaaca ggtccattat 6301 ccggttttta gtttcgctgc acattgccga atgacaatcg gattccttta agctttgctt 6361 ttgccctgtt ttctctcccc agtaatcgtt gggtgactga accatccagt agccgagagt 6421 tacgtttttc ctgatttaat aaatatccat atctatttgt ctcgtggtgt tttacagcaa 6481 ctaaccgaaa ttactatttt gcatcatcca accataaaga gacaacaata attctagaaa 6541 acatataaca agtgtttttt tctatttacg tctcatacgc aaagtccact ttaccatgtt 6601 ttggaaactt gtacccaacc attcattttt cgattttgta atgaggctct tttcttctaa 6661 caatcattga ggtcattatc atgtgtcctc acctattgtt taaattgagg cttgcgctat 6721 tcccaaaatt gtgggtatta ttttcatata tcagtatctg caatttcggt tcaactttct 6781 tatcaagaaa gaggattaat ttcttaacaa aaaccttctt tgttttataa cgctgatttt 6841 tgtattattt ggcgaccaag gttataagtg atccaatcca aatgtcataa cttcagttga 6901 tccctgcgga aaatataaat cagattatgg ttatataaaa aaaaagagaa attatttgag 6961 ttcattgaaa cttcaacgtt atttgactaa aacttgttaa tttacatgtc tgcaagatta 7021 atccagcata ttaaataaat tagttgactt atgtcaaaca aataatacat tcttgatatt 7081 attagaaaaa aaaaagaaaa ttgaataact tgtgagaaat gatattgtta ttattttaga 7141 ttatgtcctt tgaaatacaa taacatgcta aggcacttac tacttaagcc tttattactt 7201 taatgtgcta acaatcttct gtatccttag acgttaaatt ttcccttatt aaaaaataaa 7261 aaaataaaaa aaaatatagg aaatgaaaca accgctgatg acttattctg tcttttatgg 7321 cgataatttt ctgatccttc cggaagctcg gtacccaact gtgggaatac tcaggtatcg 7381 taagatgcaa gagttcgaat ctcttagcaa ccattatttt ttttctctca acataacgag 7441 aacacacagg ggcgctatcg cacagaatca aattcgatga ctggaaattt tttgttaatt 7501 tcagaggtcg cctgacgcat ataccttttt caactgaaaa attgggagaa aaaggaaagg 7561 tgagagcgcc ggaaccggct tttcatatag aatagagaag cgttcatgac taaatgcttg 7621 catcacaata cttgaagttg acaatattat ttaaggacct attgtttttt ccaataggtg 7681 gttagcaatc gtcttacttt ctaacttttc ttacctttta catttcagca atatatatat 7741 atatatatat atatatttca aggatatacc attgtaatgt ctgcccctaa gaagatcgtc 7801 gttttgccag gtgaccacgt tggtcaagaa atcacagccg aagccattaa ggttcttaaa 7861 gctatttctg atgttcgttc caatgtcaag ttcgatttcg aaaatcattt aattggtggt 7921 gctgctatcg atgctacagg tgttccactt ccagatgagg cgctggaagc ctccaagaag 7981 gctgatgccg ttttgttagg tgctgtgggt ggtcctaaat ggggtaccgg tagtgttaga 8041 cctgaacaag gtttactaaa aatccgtaaa gaacttcaat tgtacgccaa cttaagacca 8101 tgtaactttg catccgactc tcttttagac ttatctccaa tcaagccaca atttgctaaa 8161 ggtactgact tcgttgttgt tagagaatta gtgggaggta tttactttgg taagagaaag 8221 gaagacgatg gtgatggtgt cgcttgggat agtgaacaat acaccgttcc agaagtgcaa 8281 agaatcacaa gaatggccgc tttcatggcc ctacaacatg agccaccatt gcctatttgg 8341 tccttggata aagctaatgt tttggcctct tcaagattat ggagaaaaac tgtggaggaa 8401 accatcaaga acgaatttcc tacattgaaa gttcaacatc aattgattga ttctgccgcc 8461 atgatcctag ttaagaaccc aacccaccta aatggtatta taatcaccag caacatgttt 8521 ggtgatatca tctccgatga agcctccgtt atcccaggct ccttgggttt gttgccatct 8581 gcgtccttgg cctctttgcc agacaagaac accgcatttg gtttgtacga accatgccat 8641 ggttccgctc cagatttgcc aaagaataag gtcaacccta tcgccactat cttgtctgct 8701 gcaatgatgt tgaaattgtc attgaacttg cctgaagaag gtaaagccat tgaagatgca 8761 gttaaaaagg ttttggatgc aggtatcaga actggtgatt taggtggttc caacagtacc 8821 accgaagtcg gtgatgctgt cgccgaagaa gttaagaaaa tccttgctta aaaagattct 8881 ctttttttgt gatatttgta cataaacttt ataaatgaaa ttcataatag aaacgacacg 8941 aaattacaaa atggaatatg ttcatagggt agacgaaact atatacgcaa tctacataca 9001 tttatcaaga aggagaaaaa ggaggatgta aaggaataca ggtaagcaaa ttgatactaa 9061 tggctcaacg tgataaggaa aaagaattgc actttaacat taatattgac aaggaggagg 9121 gcatcaca //