LOCUS       sequence-233957-        9128 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre-expression plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
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 Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Gene."; 
            date: "2008-01-15"; volume: "407(1-2)"; pages: "63-7"
COMMENT     Sequence Label: pAW5
FEATURES             Location/Qualifiers
     source          1..9128
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     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             join(1821..1889,1890..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        join(3764..3769,3770..3787,3788..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"
     promoter        complement(6170..7323)
                     /label=nmt1 P41 promoter
                     /note="mutant nmt1 promoter from Schizosaccharomyces pombe,
                     conferring medium strength thiamine-repressible expression"
     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
//
