LOCUS       sequence-120253-        9645 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  co-expressing Cas9 and Cre recombinase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9645)
  AUTHORS   Chen Y, Cao J, Xiong M, Petersen AJ, Dong Y, Tao Y, Huang CT, Du Z, 
            Zhang SC
  TITLE     Engineering Human Stem Cell Lines with Inducible Gene Knockout using
            CRISPR/Cas9.
  JOURNAL   Cell Stem Cell. 2015 Jul 1. pii: S1934-5909(15)00261-1. doi: 
            10.1016/j.stem.2015.06.001.
  PUBMED    26145478
REFERENCE   2  (bases 1 to 9645)
  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: "Cell Stem 
            Cell. 2015 Jul 1. pii: S1934-5909(15)00261-1. doi: 
            10.1016/j.stem.2015.06.001."
COMMENT     Sequence Label: Cas9-2A-Cre
FEATURES             Location/Qualifiers
     source          1..9645
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        390..665
                     /label=chicken beta-actin promoter
     intron          666..1678
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1602..1624
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1686..1705
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1739..1748
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1745..5848
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /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"
     CDS             1745..5848
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /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"
     CDS             1745..5848
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /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"
     CDS             5861..5881
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5897..5953
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5960..6988
                     /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="MANLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFSEHTWKML
                     LSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNMLHRRSGL
                     PRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQDIRNLAF
                     LGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSLGVTKLVE
                     RWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYGAKDDSGQ
                     RYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETGAMVRLLE
                     DGD"
     CDS             6998..7027
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     polyA_signal    7087..7208
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7124..7143)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7178..7197
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7257..7273)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7257..7273)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7270..7292)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7281..7297
                     /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)."
     promoter        complement(join(7305..7311,7312..7329,7330..7335))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7350..7371
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7488..7505)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7659..8247)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7739..7758)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8418..9209,9210..9278))
                     /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     9041..9060
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9279..9383)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(9462..9481)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 acgcgttgac attgattatt gactagttat taatagtaat caattacggg gtcattagtt
       61 catagcccat atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga
      121 ccgcccaacg acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca
      181 atagggactt tccattgacg tcaatgggtg gactatttac ggtaaactgc ccacttggca
      241 gtacatcaag tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg
      301 cccgcctggc attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc
      361 tacgtattag tcatcgctat taccatgggt cgaggtgagc cccacgttct gcttcactct
      421 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      481 tgcagcgatg ggggcggggg gggggggggc gcgcgccagg cggggcgggg cggggcgagg
      541 ggcggggcgg ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa
      601 agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc
      661 gggcgggagt cgctgcgttg ccttcgcccc gtgccccgct ccgcgccgcc tcgcgccgcc
      721 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
      781 ctccgggctg taattagcgc ttggtttaat gacggctcgt ttcttttctg tggctgcgtg
      841 aaagccttaa agggctccgg gagggccctt tgtgcggggg ggagcggctc ggggggtgcg
      901 tgcgtgtgtg tgtgcgtggg gagcgccgcg tgcggcccgc gctgcccggc ggctgtgagc
      961 gctgcgggcg cggcgcgggg ctttgtgcgc tccgcgtgtg cgcgagggga gcgcggccgg
     1021 gggcggtgcc ccgcggtgcg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg
     1081 tgcgtggggg ggtgagcagg gggtgtgggc gcggcggtcg ggctgtaacc cccccctgca
     1141 cccccctccc cgagttgctg agcacggccc ggcttcgggt gcggggctcc gtacggggcg
     1201 tggcgcgggg ctcgccgtgc cgggcggggg gtggcggcag gtgggggtgc cgggcggggc
     1261 ggggccgcct cgggccgggg agggctcggg ggaggggcgc ggcggccccc ggagcgccgg
     1321 cggctgtcga ggcgcggcga gccgcagcca ttgcctttta tggtaatcgt gcgagagggc
     1381 gcagggactt cctttgtccc aaatctgtgc ggagccgaaa tctgggaggc gccgccgcac
     1441 cccctctagc gggcgcgggg cgaagcggtg cggcgccggc aggaaggaaa tgggcgggga
     1501 gggccttcgt gcgtcgccgc gccgccgtcc ccttctccat ctccagcctc ggggctgtcc
     1561 gcagggggac ggctgccttc gggggggacg gggcagggcg gggttcggct tctggcgtgt
     1621 gaccggcggc tctagtgcct ctgctaacca tgttcatgcc ttcttctttt tcctacagct
     1681 cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt ggcaaagaat tcgcggccgc
     1741 caccatggac aagaagtact ccattgggct cgatatcggc acaaacagcg tcggctgggc
     1801 cgtcattacg gacgagtaca aggtgccgag caaaaaattc aaagttctgg gcaataccga
     1861 tcgccacagc ataaagaaga acctcattgg cgccctcctg ttcgactccg gggagacggc
     1921 cgaagccacg cggctcaaaa gaacagcacg gcgcagatat acccgcagaa agaatcggat
     1981 ctgctacctg caggagatct ttagtaatga gatggctaag gtggatgact ctttcttcca
     2041 taggctggag gagtcctttt tggtggagga ggataaaaag cacgagcgcc acccaatctt
     2101 tggcaatatc gtggacgagg tggcgtacca tgaaaagtac ccaaccatat atcatctgag
     2161 gaagaagctt gtagacagta ctgataaggc tgacttgcgg ttgatctatc tcgcgctggc
     2221 gcatatgatc aaatttcggg gacacttcct catcgagggg gacctgaacc cagacaacag
     2281 cgatgtcgac aaactcttta tccaactggt tcagacttac aatcagcttt tcgaagagaa
     2341 cccgatcaac gcatccggag ttgacgccaa agcaatcctg agcgctaggc tgtccaaatc
     2401 ccggcggctc gaaaacctca tcgcacagct ccctggggag aagaagaacg gcctgtttgg
     2461 taatcttatc gccctgtcac tcgggctgac ccccaacttt aaatctaact tcgacctggc
     2521 cgaagatgcc aagcttcaac tgagcaaaga cacctacgat gatgatctcg acaatctgct
     2581 ggcccagatc ggcgaccagt acgcagacct ttttttggcg gcaaagaacc tgtcagacgc
     2641 cattctgctg agtgatattc tgcgagtgaa cacggagatc accaaagctc cgctgagcgc
     2701 tagtatgatc aagcgctatg atgagcacca ccaagacttg actttgctga aggcccttgt
     2761 cagacagcaa ctgcctgaga agtacaagga aattttcttc gatcagtcta aaaatggcta
     2821 cgccggatac attgacggcg gagcaagcca ggaggaattt tacaaattta ttaagcccat
     2881 cttggaaaaa atggacggca ccgaggagct gctggtaaag cttaacagag aagatctgtt
     2941 gcgcaaacag cgcactttcg acaatggaag catcccccac cagattcacc tgggcgaact
     3001 gcacgctatc ctcaggcggc aagaggattt ctaccccttt ttgaaagata acagggaaaa
     3061 gattgagaaa atcctcacat ttcggatacc ctactatgta ggccccctcg cccggggaaa
     3121 ttccagattc gcgtggatga ctcgcaaatc agaagagacc atcactccct ggaacttcga
     3181 ggaagtcgtg gataaggggg cctctgccca gtccttcatc gaaaggatga ctaactttga
     3241 taaaaatctg cctaacgaaa aggtgcttcc taaacactct ctgctgtacg agtacttcac
     3301 agtttataac gagctcacca aggtcaaata cgtcacagaa gggatgagaa agccagcatt
     3361 cctgtctgga gagcagaaga aagctatcgt ggacctcctc ttcaagacga accggaaagt
     3421 taccgtgaaa cagctcaaag aagactattt caaaaagatt gaatgtttcg actctgttga
     3481 aatcagcgga gtggaggatc gcttcaacgc atccctggga acgtatcacg atctcctgaa
     3541 aatcattaaa gacaaggact tcctggacaa tgaggagaac gaggacattc ttgaggacat
     3601 tgtcctcacc cttacgttgt ttgaagatag ggagatgatt gaagaacgct tgaaaactta
     3661 cgctcatctc ttcgacgaca aagtcatgaa acagctcaag aggcgccgat atacaggatg
     3721 ggggcggctg tcaagaaaac tgatcaatgg gatccgagac aagcagagtg gaaagacaat
     3781 cctggatttt cttaagtccg atggatttgc caaccggaac ttcatgcagt tgatccatga
     3841 tgactctctc acctttaagg aggacatcca gaaagcacaa gtttctggcc agggggacag
     3901 tcttcacgag cacatcgcta atcttgcagg tagcccagct atcaaaaagg gaatactgca
     3961 gaccgttaag gtcgtggatg aactcgtcaa agtaatggga aggcataagc ccgagaatat
     4021 cgttatcgag atggcccgag agaaccaaac tacccagaag ggacagaaga acagtaggga
     4081 aaggatgaag aggattgaag agggtataaa agaactgggg tcccaaatcc ttaaggaaca
     4141 cccagttgaa aacacccagc ttcagaatga gaagctctac ctgtactacc tgcagaacgg
     4201 cagggacatg tacgtggatc aggaactgga catcaatcgg ctctccgact acgacgtgga
     4261 tcatatcgtg ccccagtctt ttctcaaaga tgattctatt gataataaag tgttgacaag
     4321 atccgataaa aatagaggga agagtgataa cgtcccctca gaagaagttg tcaagaaaat
     4381 gaaaaattat tggcggcagc tgctgaacgc caaactgatc acacaacgga agttcgataa
     4441 tctgactaag gctgaacgag gtggcctgtc tgagttggat aaagccggct tcatcaaaag
     4501 gcagcttgtt gagacacgcc agatcaccaa gcacgtggcc caaattctcg attcacgcat
     4561 gaacaccaag tacgatgaaa atgacaaact gattcgagag gtgaaagtta ttactctgaa
     4621 gtctaagctg gtctcagatt tcagaaagga ctttcagttt tataaggtga gagagatcaa
     4681 caattaccac catgcgcatg atgcctacct gaatgcagtg gtaggcactg cacttatcaa
     4741 aaaatatccc aagcttgaat ctgaatttgt ttacggagac tataaagtgt acgatgttag
     4801 gaaaatgatc gcaaagtctg agcaggaaat aggcaaggcc accgctaagt acttctttta
     4861 cagcaatatt atgaattttt tcaagaccga gattacactg gccaatggag agattcggaa
     4921 gcgaccactt atcgaaacaa acggagaaac aggagaaatc gtgtgggaca agggtaggga
     4981 tttcgcgaca gtccggaagg tcctgtccat gccgcaggtg aacatcgtta aaaagaccga
     5041 agtacagacc ggaggcttct ccaaggaaag tatcctcccg aaaaggaaca gcgacaagct
     5101 gatcgcacgc aaaaaagatt gggaccccaa gaaatacggc ggattcgatt ctcctacagt
     5161 cgcttacagt gtactggttg tggccaaagt ggagaaaggg aagtctaaaa aactcaaaag
     5221 cgtcaaggaa ctgctgggca tcacaatcat ggagcgatca agcttcgaaa aaaaccccat
     5281 cgactttctc gaggcgaaag gatataaaga ggtcaaaaaa gacctcatca ttaagcttcc
     5341 caagtactct ctctttgagc ttgaaaacgg ccggaaacga atgctcgcta gtgcgggcga
     5401 gctgcagaaa ggtaacgagc tggcactgcc ctctaaatac gttaatttct tgtatctggc
     5461 cagccactat gaaaagctca aagggtctcc cgaagataat gagcagaagc agctgttcgt
     5521 ggaacaacac aaacactacc ttgatgagat catcgagcaa ataagcgaat tctccaaaag
     5581 agtgatcctc gccgacgcta acctcgataa ggtgctttct gcttacaata agcacaggga
     5641 taagcccatc agggagcagg cagaaaacat tatccacttg tttactctga ccaacttggg
     5701 cgcgcctgca gccttcaagt acttcgacac caccatagac agaaagcggt acacctctac
     5761 aaaggaggtc ctggacgcca cactgattca tcagtcaatt acggggctct atgaaacaag
     5821 aatcgacctc tctcagctcg gtggagacag cagggctgac cccaagaaga agaggaaggt
     5881 gaggtccgga agcggagcta ctaacttcag cctgctgaag caggctggag acgtggagga
     5941 gaaccctgga cctgctagca tggccaattt actgaccgta caccaaaatt tgcctgcatt
     6001 accggtcgat gcaacgagtg atgaggttcg caagaacctg atggacatgt tcagggatcg
     6061 ccaggcgttt tctgagcata cctggaaaat gcttctgtcc gtttgccggt cgtgggcggc
     6121 atggtgcaag ttgaataacc ggaaatggtt tcccgcagaa cctgaagatg ttcgcgatta
     6181 tcttctatat cttcaggcgc gcggtctggc agtaaaaact atccagcaac atttgggcca
     6241 gctaaacatg cttcatcgtc ggtccgggct gccacgacca agtgacagca atgctgtttc
     6301 actggttatg cggcggatcc gaaaagaaaa cgttgatgcc ggtgaacgtg caaaacaggc
     6361 tctagcgttc gaacgcactg atttcgacca ggttcgttca ctcatggaaa atagcgatcg
     6421 ctgccaggat atacgtaatc tggcatttct ggggattgct tataacaccc tgttacgtat
     6481 agccgaaatt gccaggatca gggttaaaga tatctcacgt actgacggtg ggagaatgtt
     6541 aatccatatt ggcagaacga aaacgctggt tagcaccgca ggtgtagaga aggcacttag
     6601 cctgggggta actaaactgg tcgagcgatg gatttccgtc tctggtgtag ctgatgatcc
     6661 gaataactac ctgttttgcc gggtcagaaa aaatggtgtt gccgcgccat ctgccaccag
     6721 ccagctatca actcgcgccc tggaagggat ttttgaagca actcatcgat tgatttacgg
     6781 cgctaaggat gactctggtc agagatacct ggcctggtct ggacacagtg cccgtgtcgg
     6841 agccgcgcga gatatggccc gcgctggagt ttcaataccg gagatcatgc aagctggtgg
     6901 ctggaccaat gtaaatattg tcatgaacta tatccgtaac ctggatagtg aaacaggggc
     6961 aatggtgcgc ctgctggaag atggcgatgg accggtggaa caaaaactta tttctgaaga
     7021 agatctgtga tagtgtacaa gtaactgcag cgcggggatc tcatgctgga gttcttcgcc
     7081 caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat
     7141 ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat
     7201 gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca
     7261 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     7321 agcataaagt gtaaagccta gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     7381 cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc
     7441 caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac
     7501 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     7561 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     7621 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     7681 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     7741 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     7801 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     7861 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     7921 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     7981 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     8041 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga
     8101 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     8161 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     8221 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     8281 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     8341 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     8401 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     8461 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     8521 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     8581 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     8641 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     8701 gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg
     8761 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     8821 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     8881 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     8941 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     9001 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     9061 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     9121 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     9181 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     9241 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat
     9301 tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa
     9361 aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga
     9421 tcgggagatc gatctcccga tcccctaggg tcgactctca gtacaatctg ctctgatgcc
     9481 gcatagttaa gccagtatct gctccctgct tgtgtgttgg aggtcgctga gtagtgcgcg
     9541 agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa gaatctgctt
     9601 agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatat
//
