LOCUS       sequence-171434-       12701 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  Cas9 expression vector for targeting to the AAVS1 locus.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 12701)
  AUTHORS   Bertero A, Pawlowski M, Ortmann D, Snijders K, Yiangou L, Cardoso de
            Brito M, Brown S, Bernard WG, Cooper JD, Giacomelli E, Gambardella 
            L, Hannan NR, Iyer D, Sampaziotis F, Serrano F, Zonneveld MC, Sinha 
            S, Kotter M, Vallier L
  TITLE     Optimized inducible shRNA and CRISPR/Cas9 platforms for in vitro 
            studies of human development using hPSCs.
  JOURNAL   Development. 2016 Dec 1;143(23):4405-4418.
  PUBMED    27899508
REFERENCE   2  (bases 1 to 12701)
  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: 
            "Development."; date: "2016-12-1"; volume: "143(23)"; pages: 
            "4405-4418"
COMMENT     Sequence Label: pAAV-Neo_CAG-Cas9
FEATURES             Location/Qualifiers
     source          1..12701
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     255..277
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     339..358
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      387..396
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             396..461
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             468..488
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             513..4613
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             513..4613
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             4614..4661
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(4749..4768)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    4814..4869
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(4868..4887)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(5230..5246)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    5254..5270
                     /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(5278..5284,5285..5302,5303..5308))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5323..5344
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     misc_feature    5418..6254
                     /label=HA-R
                     /note="right homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     primer_bind     complement(6288..6307)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6289..6307)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(6314..6331)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6314..6330)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6323..6345)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             6468..6770
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     6649..6668
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             complement(join(8167..8958,8959..9027))
                     /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     8790..8809
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9151..9739
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9640..9659
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9893..9910
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    10027..10048
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(10063..10068,10069..10086,10087..10093)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10101..10117
                     /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     10125..10141
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        10162..10180
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    10215..11018
                     /label=HA-L
                     /note="left homology arm from the adeno-associated virus 
                     integration site (AAVS1) within intron 1 of the human 
                     PPP1R12C gene"
     misc_feature    11025..11050
                     /label=SA
                     /note="splice acceptor site"
     CDS             11074..11127
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             11137..11940
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(11972..11989)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    11978..12202
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     enhancer        12208..12587
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ccattgcctt ttatggtaat cgtgcgagag ggcgcaggga cttcctttgt cccaaatctg
       61 tgcggagccg aaatctggga ggcgccgccg caccccctct agcgggcgcg gggcgaagcg
      121 gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt cgtgcgtcgc cgcgccgccg
      181 tccccttctc cctctccagc ctcggggctg tccgcggggg gacggctgcc ttcggggggg
      241 acggggcagg gcggggttcg gcttctggcg tgtgaccggc ggctctagag cctctgctaa
      301 ccatgttcat gccttcttct ttttcctaca gctcctgggc aacgtgctgg ttattgtgct
      361 gtctcatcat tttggcaaag aattccgcca ccatggacta taaggaccac gacggagact
      421 acaaggatca tgatattgat tacaaagacg atgacgataa gatggcccca aagaagaagc
      481 ggaaggtcgg tatccacgga gtcccagcag ccgacaagaa gtacagcatc ggcctggaca
      541 tcggcaccaa ctctgtgggc tgggccgtga tcaccgacga gtacaaggtg cccagcaaga
      601 aattcaaggt gctgggcaac accgaccggc acagcatcaa gaagaacctg atcggagccc
      661 tgctgttcga cagcggcgaa acagccgagg ccacccggct gaagagaacc gccagaagaa
      721 gatacaccag acggaagaac cggatctgct atctgcaaga gatcttcagc aacgagatgg
      781 ccaaggtgga cgacagcttc ttccacagac tggaagagtc cttcctggtg gaagaggata
      841 agaagcacga gcggcacccc atcttcggca acatcgtgga cgaggtggcc taccacgaga
      901 agtaccccac catctaccac ctgagaaaga aactggtgga cagcaccgac aaggccgacc
      961 tgcggctgat ctatctggcc ctggcccaca tgatcaagtt ccggggccac ttcctgatcg
     1021 agggcgacct gaaccccgac aacagcgacg tggacaagct gttcatccag ctggtgcaga
     1081 cctacaacca gctgttcgag gaaaacccca tcaacgccag cggcgtggac gccaaggcca
     1141 tcctgtctgc cagactgagc aagagcagac ggctggaaaa tctgatcgcc cagctgcccg
     1201 gcgagaagaa gaatggcctg ttcggaaacc tgattgccct gagcctgggc ctgaccccca
     1261 acttcaagag caacttcgac ctggccgagg atgccaaact gcagctgagc aaggacacct
     1321 acgacgacga cctggacaac ctgctggccc agatcggcga ccagtacgcc gacctgtttc
     1381 tggccgccaa gaacctgtcc gacgccatcc tgctgagcga catcctgaga gtgaacaccg
     1441 agatcaccaa ggcccccctg agcgcctcta tgatcaagag atacgacgag caccaccagg
     1501 acctgaccct gctgaaagct ctcgtgcggc agcagctgcc tgagaagtac aaagagattt
     1561 tcttcgacca gagcaagaac ggctacgccg gctacattga cggcggagcc agccaggaag
     1621 agttctacaa gttcatcaag cccatcctgg aaaagatgga cggcaccgag gaactgctcg
     1681 tgaagctgaa cagagaggac ctgctgcgga agcagcggac cttcgacaac ggcagcatcc
     1741 cccaccagat ccacctggga gagctgcacg ccattctgcg gcggcaggaa gatttttacc
     1801 cattcctgaa ggacaaccgg gaaaagatcg agaagatcct gaccttccgc atcccctact
     1861 acgtgggccc tctggccagg ggaaacagca gattcgcctg gatgaccaga aagagcgagg
     1921 aaaccatcac cccctggaac ttcgaggaag tggtggacaa gggcgcttcc gcccagagct
     1981 tcatcgagcg gatgaccaac ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc
     2041 acagcctgct gtacgagtac ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga
     2101 ccgagggaat gagaaagccc gccttcctga gcggcgagca gaaaaaggcc atcgtggacc
     2161 tgctgttcaa gaccaaccgg aaagtgaccg tgaagcagct gaaagaggac tacttcaaga
     2221 aaatcgagtg cttcgactcc gtggaaatct ccggcgtgga agatcggttc aacgcctccc
     2281 tgggcacata ccacgatctg ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg
     2341 aaaacgagga cattctggaa gatatcgtgc tgaccctgac actgtttgag gacagagaga
     2401 tgatcgagga acggctgaaa acctatgccc acctgttcga cgacaaagtg atgaagcagc
     2461 tgaagcggcg gagatacacc ggctggggca ggctgagccg gaagctgatc aacggcatcc
     2521 gggacaagca gtccggcaag acaatcctgg atttcctgaa gtccgacggc ttcgccaaca
     2581 gaaacttcat gcagctgatc cacgacgaca gcctgacctt taaagaggac atccagaaag
     2641 cccaggtgtc cggccagggc gatagcctgc acgagcacat tgccaatctg gccggcagcc
     2701 ccgccattaa gaagggcatc ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga
     2761 tgggccggca caagcccgag aacatcgtga tcgaaatggc cagagagaac cagaccaccc
     2821 agaagggaca gaagaacagc cgcgagagaa tgaagcggat cgaagagggc atcaaagagc
     2881 tgggcagcca gatcctgaaa gaacaccccg tggaaaacac ccagctgcag aacgagaagc
     2941 tgtacctgta ctacctgcag aatgggcggg atatgtacgt ggaccaggaa ctggacatca
     3001 accggctgtc cgactacgat gtggaccata tcgtgcctca gagctttctg aaggacgact
     3061 ccatcgacaa caaggtgctg accagaagcg acaagaaccg gggcaagagc gacaacgtgc
     3121 cctccgaaga ggtcgtgaag aagatgaaga actactggcg gcagctgctg aacgccaagc
     3181 tgattaccca gagaaagttc gacaatctga ccaaggccga gagaggcggc ctgagcgaac
     3241 tggataaggc cggcttcatc aagagacagc tggtggaaac ccggcagatc acaaagcacg
     3301 tggcacagat cctggactcc cggatgaaca ctaagtacga cgagaatgac aagctgatcc
     3361 gggaagtgaa agtgatcacc ctgaagtcca agctggtgtc cgatttccgg aaggatttcc
     3421 agttttacaa agtgcgcgag atcaacaact accaccacgc ccacgacgcc tacctgaacg
     3481 ccgtcgtggg aaccgccctg atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg
     3541 gcgactacaa ggtgtacgac gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca
     3601 aggctaccgc caagtacttc ttctacagca acatcatgaa ctttttcaag accgagatta
     3661 ccctggccaa cggcgagatc cggaagcggc ctctgatcga gacaaacggc gaaaccgggg
     3721 agatcgtgtg ggataagggc cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc
     3781 aagtgaatat cgtgaaaaag accgaggtgc agacaggcgg cttcagcaaa gagtctatcc
     3841 tgcccaagag gaacagcgat aagctgatcg ccagaaagaa ggactgggac cctaagaagt
     3901 acggcggctt cgacagcccc accgtggcct attctgtgct ggtggtggcc aaagtggaaa
     3961 agggcaagtc caagaaactg aagagtgtga aagagctgct ggggatcacc atcatggaaa
     4021 gaagcagctt cgagaagaat cccatcgact ttctggaagc caagggctac aaagaagtga
     4081 aaaaggacct gatcatcaag ctgcctaagt actccctgtt cgagctggaa aacggccgga
     4141 agagaatgct ggcctctgcc ggcgaactgc agaagggaaa cgaactggcc ctgccctcca
     4201 aatatgtgaa cttcctgtac ctggccagcc actatgagaa gctgaagggc tcccccgagg
     4261 ataatgagca gaaacagctg tttgtggaac agcacaagca ctacctggac gagatcatcg
     4321 agcagatcag cgagttctcc aagagagtga tcctggccga cgctaatctg gacaaagtgc
     4381 tgtccgccta caacaagcac cgggataagc ccatcagaga gcaggccgag aatatcatcc
     4441 acctgtttac cctgaccaat ctgggagccc ctgccgcctt caagtacttt gacaccacca
     4501 tcgaccggaa gaggtacacc agcaccaaag aggtgctgga cgccaccctg atccaccaga
     4561 gcatcaccgg cctgtacgag acacggatcg acctgtctca gctgggaggc gacaaaaggc
     4621 cggcggccac gaaaaaggcc ggccaggcaa aaaagaaaaa gtaatgaatt cactcctcag
     4681 gtgcaggctg cctatcagaa ggtggtggct ggtgtggcca atgccctggc tcacaaatac
     4741 cactgagatc tttttccctc tgccaaaaat tatggggaca tcatgaagcc ccttgagcat
     4801 ctgacttctg gctaataaag gaaatttatt ttcattgcaa tagtgtgttg gaattttttg
     4861 tgtctctcac tcggaaggac atatgggagg gcaaatcatt taaaacatca gaatgagtat
     4921 ttggtttaga gtttggcaac atatgcccat atgctggctg ccatgaacaa aggttggcta
     4981 taaagaggtc atcagtatat gaaacagccc cctgctgtcc attccttatt ccatagaaaa
     5041 gccttgactt gaggttagat tttttttata ttttgttttg tgttattttt ttctttaaca
     5101 tccctaaaat tttccttaca tgttttacta gccagatttt tcctcctctc ctgactactc
     5161 ccagtcatag ctgtccctct tctcttatgg agatccctcg acctgcagcc caagcttggc
     5221 gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa
     5281 catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac
     5341 attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagcggat
     5401 cgacagtact aagctttact agggacagga ttggtgacag aaaagcccca tccttaggcc
     5461 tcctccttcc tagtctcctg atattgggtc taacccccac ctcctgttag gcagattcct
     5521 tatctggtga cacaccccca tttcctggag ccatctctct ccttgccaga acctctaagg
     5581 tttgcttacg atggagccag agaggatcct gggagggaga gcttggcagg gggtgggagg
     5641 gaaggggggg atgcgtgacc tgcccggttc tcagtggcca ccctgcgcta ccctctccca
     5701 gaacctgagc tgctctgacg cggctgtctg gtgcgtttca ctgatcctgg tgctgcagct
     5761 tccttacact tcccaagagg agaagcagtt tggaaaaaca aaatcagaat aagttggtcc
     5821 tgagttctaa ctttggctct tcacctttct agtccccaat ttatattgtt cctccgtgcg
     5881 tcagttttac ctgtgagata aggccagtag ccagccccgt cctggcaggg ctgtggtgag
     5941 gaggggggtg tccgtgtgga aaactccctt tgtgagaatg gtgcgtccta ggtgttcacc
     6001 aggtcgtggc cgcctctact ccctttctct ttctccatcc ttctttcctt aaagagtccc
     6061 cagtgctatc tgggacatat tcctccgccc agagcagggt cccgcttccc taaggccctg
     6121 ctctgggctt ctgggtttga gtccttggca agcccaggag aggcgctcag gcttccctgt
     6181 cccccttcct cgtccaccat ctcatgcccc tggctctcct gccccttccc tacaggggtt
     6241 cctggctctg ctctaagggc gaattcgcgg ccgctaaatt caattcgccc tatagtgagt
     6301 cgtattacaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     6361 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     6421 cccgcaccga tcgcccttcc caacagttgc gcagcctata cgtacggcag tttaaggttt
     6481 acacctataa aagagagagc cgttatcgtc tgtttgtgga tgtacagagt gatattattg
     6541 acacgccggg gcgacggatg gtgatccccc tggccagtgc acgtctgctg tcagataaag
     6601 tctcccgtga actttacccg gtggtgcata tcggggatga aagctggcgc atgatgacca
     6661 ccgatatggc cagtgtgccg gtctccgtta tcggggaaga agtggctgat ctcagccacc
     6721 gcgaaaatga catcaaaaac gccattaacc tgatgttctg gggaatataa atgtcaggca
     6781 tgagattatc aaaaaggatc ttcacctaga tccttttcac gtagaaagcc agtccgcaga
     6841 aacggtgctg accccggatg aatgtcagct actgggctat ctggacaagg gaaaacgcaa
     6901 gcgcaaagag aaagcaggta gcttgcagtg ggcttacatg gcgatagcta gactgggcgg
     6961 ttttatggac agcaagcgaa ccggaattgc cagctggggc gccctctggt aaggttggga
     7021 agccctgcaa agtaaactgg atggctttct tgccgccaag gatctgatgg cgcaggggat
     7081 caagctctga tcaagagaca ggatgaggat cgtttcgcat gattgaacaa gatggattgc
     7141 acgcaggttc tccggccgct tgggtggaga ggctattcgg ctatgactgg gcacaacaga
     7201 caatcggctg ctctgatgcc gccgtgttcc ggctgtcagc gcaggggcgc ccggttcttt
     7261 ttgtcaagac cgacctgtcc ggtgccctga atgaactgca agacgaggca gcgcggctat
     7321 cgtggctggc cacgacgggc gttccttgcg cagctgtgct cgacgttgtc actgaagcgg
     7381 gaagggactg gctgctattg ggcgaagtgc cggggcagga tctcctgtca tctcaccttg
     7441 ctcctgccga gaaagtatcc atcatggctg atgcaatgcg gcggctgcat acgcttgatc
     7501 cggctacctg cccattcgac caccaagcga aacatcgcat cgagcgagca cgtactcgga
     7561 tggaagccgg tcttgtcgat caggatgatc tggacgaaga gcatcagggg ctcgcgccag
     7621 ccgaactgtt cgccaggctc aaggcgagca tgcccgacgg cgaggatctc gtcgtgaccc
     7681 atgcatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt ttctggattc
     7741 atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt ggctacccgt
     7801 gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc
     7861 gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt cttctgaatt
     7921 attaacgctt acaatttcct gatgcggtat tttctcctta cgcatctgtg cggtatttca
     7981 caccgcatca ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt
     8041 ctaaatacat tcaaatatgt atccgctcat gagattatca aaaaggatct tcacctagat
     8101 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     8161 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     8221 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     8281 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     8341 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     8401 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     8461 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     8521 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     8581 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     8641 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     8701 cttttctgtg actggtgaat actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     8761 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     8821 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     8881 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     8941 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     9001 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     9061 tcagggttat tgtctcatga ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc
     9121 agaccccgta gaaaagatca aaggatcttc ttgagatcct ttttttctgc gcgtaatctg
     9181 ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt tgtttgccgg atcaagagct
     9241 accaactctt tttccgaagg taactggctt cagcagagcg cagataccaa atactgttct
     9301 tctagtgtag ccgtagttag gccaccactt caagaactct gtagcaccgc ctacatacct
     9361 cgctctgcta atcctgttac cagtggctgc tgccagtggc gataagtcgt gtcttaccgg
     9421 gttggactca agacgatagt taccggataa ggcgcagcgg tcgggctgaa cggggggttc
     9481 gtgcacacag cccagcttgg agcgaacgac ctacaccgaa ctgagatacc tacagcgtga
     9541 gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg
     9601 cagggtcgga acaggagagc gcacgaggga gcttccaggg ggaaacgcct ggtatcttta
     9661 tagtcctgtc gggtttcgcc acctctgact tgagcgtcga tttttgtgat gctcgtcagg
     9721 ggggcggagc ctatggaaaa acgccagcaa cgcggccttt ttacggttcc tggccttttg
     9781 ctggcctttt gctcacatgt tctttcctgc gttatcccct gattctgtgg ataaccgtat
     9841 taccgccttt gagtgagctg ataccgctcg ccgcagccga acgaccgagc gcagcgagtc
     9901 agtgagcgag gaagcggaag agcgcccaat acgcaaaccg cctctccccg cgcgttggcc
     9961 gattcattaa tgcagctggc acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa
    10021 cgcaattaat gtgagttagc tcactcatta ggcaccccag gctttacact ttatgcttcc
    10081 ggctcgtatg ttgtgtggaa ttgtgagcgg ataacaattt cacacaggaa acagctatga
    10141 ccatgattac gccaagctca gaattaaccc tcactaaagg gactagtcct gcaggtttaa
    10201 acgaattcgc cctttgcttt ctctgaccag cattctctcc cctgggcctg tgccgctttc
    10261 tgtctgcagc ttgtggcctg ggtcacctct acggctggcc cagatccttc cctgccgcct
    10321 ccttcaggtt ccgtcttcct ccactccctc ttccccttgc tctctgctgt gttgctgccc
    10381 aaggatgctc tttccggagc acttccttct cggcgctgca ccacgtgatg tcctctgagc
    10441 ggatcctccc cgtgtctggg tcctctccgg gcatctctcc tccctcaccc aaccccatgc
    10501 cgtcttcact cgctgggttc ccttttcctt ctccttctgg ggcctgtgcc atctctcgtt
    10561 tcttaggatg gccttctccg acggatgtct cccttgcgtc ccgcctcccc ttcttgtagg
    10621 cctgcatcat caccgttttt ctggacaacc ccaaagtacc ccgtctccct ggctttagcc
    10681 acctctccat cctcttgctt tctttgcctg gacaccccgt tctcctgtgg attcgggtca
    10741 cctctcactc ctttcatttg ggcagctccc ctacccccct tacctctcta gtctgtgcta
    10801 gctcttccag ccccctgtca tggcatcttc caggggtccg agagctcagc tagtcttctt
    10861 cctccaaccc gggcccctat gtccacttca ggacagcatg tttgctgcct ccagggatcc
    10921 tgtgtccccg agctgggacc accttatatt cccagggccg gttaatgtgg ctctggttct
    10981 gggtactttt atctgtcccc tccaccccac agtggggcaa gcttctgacc tcttctcttc
    11041 ctcccacagg gcctcgagag atctggcagc ggagagggca gaggaagtct tctaacatgc
    11101 ggtgacgtgg aggagaatcc cggccctagg ctcgagatgg gatcggccat tgaacaagat
    11161 ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca
    11221 caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg
    11281 gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga cgaggcagcg
    11341 cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact
    11401 gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct
    11461 caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg
    11521 cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt
    11581 actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc
    11641 gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga tgatctcgtc
    11701 gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga
    11761 ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc
    11821 cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt
    11881 atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga
    11941 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
    12001 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
    12061 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
    12121 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
    12181 ggggatgcgg tgggctctat gggtctcgac attgattatt gactagttat taatagtaat
    12241 caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca taacttacgg
    12301 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
    12361 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
    12421 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
    12481 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
    12541 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc gaggtgagcc
    12601 ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat
    12661 ttatttttta attattttgt gcagcgatgg gggcgggggg g
//
