LOCUS       pAW8-yEGFP.       10655 bp DNA     circular SYN 30-NOV-2018
DEFINITION  Cloning vector pAW8-yEGFP, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    pAW8-yEGFP.
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10655)
  AUTHORS   Watson AT, Garcia V, Bone N, Carr AM, Armstrong J.
  TITLE     Gene tagging and gene replacement using recombinase-mediated 
            cassette exchange in Schizosaccharomyces pombe
  JOURNAL   Gene 407 (1-2), 63-74 (2008)
  PUBMED    18054176
REFERENCE   2  (bases 1 to 10655)
  AUTHORS   Watson AT.
  TITLE     Direct Submission
  JOURNAL   Submitted (13-AUG-2007) Genome Damage and Stability Centre, 
            University of Sussex, Falmer, Brighton, East Sussex BN1 9RQ, UK
REFERENCE   3  (bases 1 to 10655)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 10655)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Gene 407 
            (1-2), 63-74 (2008)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (13-AUG-2007) Genome Damage and Stability Centre, University of 
            Sussex, Falmer, Brighton, East Sussex BN1 9RQ, UK"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..10655
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      575..1356
                     /label=ars1
                     /note="Schizosaccharomyces pombe autonomously replicating 
                     sequence ars1"
     promoter        1716..1820
                     /label=AmpR promoter
     CDS             1821..2678
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      2852..3440
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    3728..3749
                     /label=CAP binding site
                     /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 repressor encoded by lacI binding site
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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)."
     primer_bind     3826..3842
                     /label=M13 rev
                     /note="M13 rev"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     gene            complement(3976..5332)
                     /label=kanMX
                     /note="yeast selectable marker conferring kanamycin
                     resistance (Wach et al., 1994)"
     terminator      complement(5407..5594)
                     /label=ADH1 terminator
                     /note="transcription terminator for the S. cerevisiae
                     alcohol dehydrogenase 1 (ADH1) gene"
     CDS             complement(5618..6331)
                     /label=GFP
                     /note="green fluorescent protein"
     protein_bind    6340..6373
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(6389..6405)
                     /label=M13 fwd
                     /note="M13 fwd"
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      6612..7168
                     /label=nmt1
                     /note="nmt1"
                     /regulatory_class="terminator"
     CDS             complement(7177..8166)
                     /label=Cre
                     /note="site-specific recombinase"
     intron          complement(8167..8215)
                     /gene="cre"
                     /note="Schizosaccharomyces pombe rad50 intron 1"
     regulatory      complement(8255..8865)
                     /label=nmt41
                     /note="nmt41"
                     /regulatory_class="promoter"
     promoter        8884..9303
                     /label=LEU2 promoter
     CDS             9304..10395
                     /label=LEU2
                     /note="3-isopropylmalate dehydrogenase, required for
                     leucine biosynthesis"
     regulatory      10399..10655
                     /gene="LEU2"
                     /regulatory_class="terminator"
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 ggtgtagatt 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 gtcaggccac 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 gcatgcgcgg ccgcataact tcgtatataa taccatatac
     3901 gaagttatac tagttcgtcg acggtatcga taagcttgat tcgactggat ggcggcgtta
     3961 gtatcgaatc gacagcagta tagcgaccag cattcacata cgattgacgc atgatattac
     4021 tttctgcgca cttaacttcg catctgggca gatgatgtcg aggcgaaaaa aaatataaat
     4081 cacgctaaca tttgattaaa atagaacaac tacaatataa aaaaactata caaatgacaa
     4141 gttcttgaaa acaagaatct ttttattgtc agtactgatt agaaaaactc atcgagcatc
     4201 aaatgaaact gcaatttatt catatcagga ttatcaatac catatttttg aaaaagccgt
     4261 ttctgtaatg aaggagaaaa ctcaccgagg cagttccata ggatggcaag atcctggtat
     4321 cggtctgcga ttccgactcg tccaacatca atacaaccta ttaatttccc ctcgtcaaaa
     4381 ataaggttat caagtgagaa atcaccatga gtgacgactg aatccggtga gaatggcaaa
     4441 agcttatgca tttctttcca gacttgttca acaggccagc cattacgctc gtcatcaaaa
     4501 tcactcgcat caaccaaacc gttattcatt cgtgattgcg cctgagcgag acgaaatacg
     4561 cgatcgctgt taaaaggaca attacaaaca ggaatcgaat gcaaccggcg caggaacact
     4621 gccagcgcat caacaatatt ttcacctgaa tcaggatatt cttctaatac ctggaatgct
     4681 gttttgccgg ggatcgcagt ggtgagtaac catgcatcat caggagtacg gataaaatgc
     4741 ttgatggtcg gaagaggcat aaattccgtc agccagttta gtctgaccat ctcatctgta
     4801 acatcattgg caacgctacc tttgccatgt ttcagaaaca actctggcgc atcgggcttc
     4861 ccatacaatc gatagattgt cgcacctgat tgcccgacat tatcgcgagc ccatttatac
     4921 ccatataaat cagcatccat gttggaattt aatcgcggcc tcgaaacgtg agtcttttcc
     4981 ttacccatgg ttgtttatgt tcggatgtga tgtgagaact gtatcctagc aagattttaa
     5041 aaggaagtat atgaaagaag aacctcagtg gcaaatccta accttttata tttctctaca
     5101 ggggcgcggc gtggggacaa ttcaacgcgt ctgtgagggg agcgtttccc tgctcgcagg
     5161 tctgcagcga ggagccgtaa tttttgcttc gcgccgtgcg gccatcaaaa tgtatggatg
     5221 caaatgatta tacatgggga tgtatgggct aaatgtacgg gcgacagtca catcatgccc
     5281 ctgagctgcg cacgtcaaga ctgtcaagga gggtattctg ggcctccatg tcgctggccg
     5341 ggtgacccgg cggggacgag gcaagctaaa cagatctata ttaccctgtt atccctagcg
     5401 gatctgccgg tagaggtgtg gtcaataaga gcgacctcat actatacctg agaaagcaac
     5461 ctgacctaca ggaaagagtt actcaagaat aagaattttc gttttaaaac ctaagagtca
     5521 ctttaaaatt tgtatacact tatttttttt ataacttatt taataataaa aatcataaat
     5581 cataagaaat tcgcttattt agaagtggcg cgccttattt gtacaattca tccataccat
     5641 gggtaatacc agcagcagta acaaattcta acaagaccat gtggtctctc ttttcgtttg
     5701 gatctttgga taaggcagat tgagtggata agtaatggtt gtctggtaac aagactggac
     5761 catcaccaat tggagtattt tgttgataat ggtcagctaa ttgaacagaa ccatcttcaa
     5821 tgttgtgtct aattttgaag ttaactttga taccattctt ttgtttgtca gccatgatgt
     5881 aaacattgtg agagttatag ttgtattcca atttgtgacc taaaatgtta ccatcttctt
     5941 taaaatcaat accttttaat tcgattctat taactaaggt atcaccttca aacttgactt
     6001 cagctctggt cttgtagtta ccgtcatctt tgaaaaaaat agttctttct tgaacataac
     6061 cttctggcat ggcagacttg aaaaagtcat gttgtttcat atgatctggg tatctagaaa
     6121 aacattgaac accataagtt aaagtagtga ctaaggttgg ccatggaact ggcaatttac
     6181 cagtagtaca aataaatttt aaggtcaatt taccgtaagt agcatcacct tcaccttcac
     6241 cggagacaga aaatttgtga ccattaacat caccatctaa ttcaaccaaa attgggacaa
     6301 caccagtgaa taattcttca cctttagaca tgagcatgca taacttcgta taatgtatgc
     6361 tatacgaagt tatgcggccg ctgaattcac tggccgtcgt tttacaacgt cgtgactggg
     6421 aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc
     6481 gtaatagcga agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg
     6541 aatggcgcct gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac
     6601 catcgtccgg atcaatgaag tagatgagtt tactacctgt atatactttt ttttttgtca
     6661 aagtcacttt tatggcattt ccatgttgaa ggattagcca cagcactaaa ttttcagcgt
     6721 atgattgctt ttaaatattt aattttcatc gttttttaat taatattcaa aacgatttaa
     6781 tgccttctac atctgaagtc aagctcatag actcgggttg ggtagctaaa actacatcta
     6841 acacgaaggg gtatagcctt cttaaagtat actcatgtat aaacgttttg ggttcatatt
     6901 tttgggccac gtggtttata aaaattctta actacaccac tcggtatccc gcacccgtct
     6961 acgtttctac gatttcgagc taatattgag taaagtgatt agcaaaaaaa ataaaagtac
     7021 tcgttgtcgg agatcaagaa tttttctatt atctcatcta aaccactttc taaaagcgaa
     7081 aaacaaaatc gtaatatgca gcttgaatgg gcttccatag tttgaaagaa aaaccctagc
     7141 agtactggca agggagacat tccttttacc tggctaatcg ccatcttcca gcaggcgcac
     7201 cattgcccct gtttcactat ccaggttacg gatatagttc atgacaatat ttacattggt
     7261 ccagccacca gcttgcatga tctccggtat tgaaactcca gcgcgggcca tatctcgcgc
     7321 ggctccgaca cgggcactgt gtccagacca ggccaggtat ctctgaccag agtcatcctt
     7381 agcgccgtaa atcaatcgat gagttgcttc aaaaatccct tccagggcgc gagttgatag
     7441 ctggctggtg gcagatggcg cggcaacacc attttttctg acccggcaaa acaggtagtt
     7501 attcggatca tcagctacac cagagacgga aatccatcgc tcgaccagtt tagttacccc
     7561 caggctaagt gccttctcta cacctgcggt gctaaccagc gttttcgttc tgccaatatg
     7621 gattaacatt ctcccaccgt cagtacgtga gatatcttta accctgatcc tggcaatttc
     7681 ggctatacgt aacagggtgt tataagcaat ccccagaaat gccagattac gtatatcctg
     7741 gcagcgatcg ctattttcca tgagtgaacg aacctggtcg aaatcagtgc gttcgaacgc
     7801 tagagcctgt tttgcacgtt caccggcatc aacgttttct tttcggatcc gccgcataac
     7861 cagtgaaaca gcattgctgt cacttggtcg tggcagcccg gaccgacgat gaagcatgtt
     7921 tagctggccc aaatgttgct ggatagtttt tactgccaga ccgcgcgcct gaagatatag
     7981 aagataatcg cgaacatctt caggttctgc gggaaaccat ttccggttat tcaacttgca
     8041 ccatgccgcc cacgaccggc aaacggacag aagcattttc caggtatgct cagaaaacgc
     8101 ctggcgatcc ctgaacatgt ccatcaggtt cttgcgaacc tcatcactcg ttgcatcgac
     8161 cggtaacttt tagattaaca tttgtttaaa tatgaaatta ggcaatgtaa catactgcag
     8221 gcaaattttg gtgtacggta agtaaattgg acatatgatt taacaaagcg actataagtc
     8281 agaaagtgag aatgagattg aaataattaa ttcaacttat tcaattgatg atatgccagg
     8341 attcctcttc ctttatatat cttcaatgtt tcgattaaca ggtccattat ccggttttta
     8401 gtttcgctgc acattgccga atgacaatcg gattccttta agctttgctt ttgccctgtt
     8461 ttctctcccc agtaatcgtt gggtgactga accatccagt agccgagagt tacgtttttc
     8521 ctgatttaat aaatatccat atctatttgt ctcgtggtgt tttacagcaa ctaaccgaaa
     8581 ttactatttt gcatcatcca accataaaga gacaacaata attctagaaa acatataaca
     8641 agtgtttttt tctatttacg tctcatacgc aaagtccact ttaccatgtt ttggaaactt
     8701 gtacccaacc attcattttt cgattttgta atgaggctct tttcttctaa caatcattga
     8761 ggtcattatc atgtgtcctc acctattgtt taaattgagg cttgcgctat tcccaaaatt
     8821 gtgggtatta ttttcatata tcagtatctg caatttcggt tcaactccgg aagctcggta
     8881 cccaactgtg ggaatactca ggtatcgtaa gatgcaagag ttcgaatctc ttagcaacca
     8941 ttattttttt tctctcaaca taacgagaac acacaggggc gctatcgcac agaatcaaat
     9001 tcgatgactg gaaatttttt gttaatttca gaggtcgcct gacgcatata cctttttcaa
     9061 ctgaaaaatt gggagaaaaa ggaaaggtga gagcgccgga accggctttt catatagaat
     9121 agagaagcgt tcatgactaa atgcttgcat cacaatactt gaagttgaca atattattta
     9181 aggacctatt gttttttcca ataggtggtt agcaatcgtc ttactttcta acttttctta
     9241 ccttttacat ttcagcaata tatatatata tatatatata tatttcaagg atataccatt
     9301 gtaatgtctg cccctaagaa gatcgtcgtt ttgccaggtg accacgttgg tcaagaaatc
     9361 acagccgaag ccattaaggt tcttaaagct atttctgatg ttcgttccaa tgtcaagttc
     9421 gatttcgaaa atcatttaat tggtggtgct gctatcgatg ctacaggtgt tccacttcca
     9481 gatgaggcgc tggaagcctc caagaaggct gatgccgttt tgttaggtgc tgtgggtggt
     9541 cctaaatggg gtaccggtag tgttagacct gaacaaggtt tactaaaaat ccgtaaagaa
     9601 cttcaattgt acgccaactt aagaccatgt aactttgcat ccgactctct tttagactta
     9661 tctccaatca agccacaatt tgctaaaggt actgacttcg ttgttgttag agaattagtg
     9721 ggaggtattt actttggtaa gagaaaggaa gacgatggtg atggtgtcgc ttgggatagt
     9781 gaacaataca ccgttccaga agtgcaaaga atcacaagaa tggccgcttt catggcccta
     9841 caacatgagc caccattgcc tatttggtcc ttggataaag ctaatgtttt ggcctcttca
     9901 agattatgga gaaaaactgt ggaggaaacc atcaagaacg aatttcctac attgaaagtt
     9961 caacatcaat tgattgattc tgccgccatg atcctagtta agaacccaac ccacctaaat
    10021 ggtattataa tcaccagcaa catgtttggt gatatcatct ccgatgaagc ctccgttatc
    10081 ccaggctcct tgggtttgtt gccatctgcg tccttggcct ctttgccaga caagaacacc
    10141 gcatttggtt tgtacgaacc atgccatggt tccgctccag atttgccaaa gaataaggtc
    10201 aaccctatcg ccactatctt gtctgctgca atgatgttga aattgtcatt gaacttgcct
    10261 gaagaaggta aagccattga agatgcagtt aaaaaggttt tggatgcagg tatcagaact
    10321 ggtgatttag gtggttccaa cagtaccacc gaagtcggtg atgctgtcgc cgaagaagtt
    10381 aagaaaatcc ttgcttaaaa agattctctt tttttgtgat atttgtacat aaactttata
    10441 aatgaaattc ataatagaaa cgacacgaaa ttacaaaatg gaatatgttc atagggtaga
    10501 cgaaactata tacgcaatct acatacattt atcaagaagg agaaaaagga ggatgtaaag
    10561 gaatacaggt aagcaaattg atactaatgg ctcaacgtga taaggaaaaa gaattgcact
    10621 ttaacattaa tattgacaag gaggagggca tcaca
//
