LOCUS       sequence-488593-        8506 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SpCas9 and a sgRNA targeting the human histone H3.2 loci 
            for knock-in..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8506)
  AUTHORS   Minami K, Nakazato K, Ide S, Kaizu K, Higashi K, Tamura S, Toyoda A,
            Takahashi K, Kurokawa K, Maeshima K
  TITLE     Replication-dependent histone labeling dissects the physical 
            properties of euchromatin/heterochromatin in living human cells.
  JOURNAL   Sci Adv. 2025 Mar 28;11(13):eadu8400. doi: 10.1126/sciadv.adu8400. 
            Epub 2025 Mar 28.
  PUBMED    40153514
REFERENCE   2  (bases 1 to 8506)
  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: "Sci Adv."; 
            date: "2025-03-28"; volume: "11(13):eadu8400. doi"; pages: " 
            10.1126/sciadv.adu8400. Epub 2025 Mar 28"
COMMENT     Sequence Label: pX330_H3.2
FEATURES             Location/Qualifiers
     source          1..8506
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        271..346
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        443..728
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        730..1005
                     /label=chicken beta-actin promoter
     intron          1006..1233
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1245..1254
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1254..1319
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1326..1346
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1371..5471
                     /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             1371..5471
                     /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             5472..5519
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5547..5564)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5553..5760
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5769..5909
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5769..5898
                     /label=AAV2 ITR
     rep_origin      5984..6439
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6071..6090)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6281..6302
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6456..6475)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6575..6597
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6635..6653)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6721..6825
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6826..6894,6895..7686)
                     /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(7044..7063)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7857..8445
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8346..8365
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ccgccgcatc cgtggagagc gttttagagc tagaaatagc aagttaaaat
      301 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgttttaga
      361 gctagaaata gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc caattctgca
      421 gacaaatggc tctagaggta cccgttacat aacttacggt aaatggcccg cctggctgac
      481 cgcccaacga cccccgccca ttgacgtcaa tagtaacgcc aatagggact ttccattgac
      541 gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata
      601 tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattgtgccc
      661 agtacatgac cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta
      721 ttaccatggt cgaggtgagc cccacgttct gcttcactct ccccatctcc cccccctccc
      781 cacccccaat tttgtattta tttatttttt aattattttg tgcagcgatg ggggcggggg
      841 gggggggggc gcgcgccagg cggggcgggg cggggcgagg ggcggggcgg ggcgaggcgg
      901 agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa agtttccttt tatggcgagg
      961 cggcggcggc ggcggcccta taaaaagcga agcgcgcggc gggcgggagt cgctgcgcgc
     1021 tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg
     1081 accgcgttac tcccacaggt gagcgggcgg gacggccctt ctcctccggg ctgtaattag
     1141 ctgagcaaga ggtaagggtt taagggatgg ttggttggtg gggtattaat gtttaattac
     1201 ctggagcacc tgcctgaaat cacttttttt caggttggac cggtgccacc atggactata
     1261 aggaccacga cggagactac aaggatcatg atattgatta caaagacgat gacgataaga
     1321 tggccccaaa gaagaagcgg aaggtcggta tccacggagt cccagcagcc gacaagaagt
     1381 acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt
     1441 acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga
     1501 agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc acccggctga
     1561 agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga
     1621 tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct
     1681 tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg
     1741 aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca
     1801 gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc
     1861 ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt
     1921 tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg
     1981 gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc
     2041 tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg attgccctga
     2101 gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc
     2161 agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc
     2221 agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca
     2281 tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat
     2341 acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg
     2401 agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacattgacg
     2461 gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg
     2521 gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct
     2581 tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc
     2641 ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga
     2701 ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga
     2761 tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg
     2821 gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg
     2881 agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat aacgagctga
     2941 ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga
     3001 aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga
     3061 aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag
     3121 atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg
     3181 acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac
     3241 tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg
     3301 acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga
     3361 agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt
     3421 ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta
     3481 aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg
     3541 ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg
     3601 acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca
     3661 gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg
     3721 aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc
     3781 agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg
     3841 accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc gtgcctcaga
     3901 gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac aagaaccggg
     3961 gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcggc
     4021 agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc aaggccgaga
     4081 gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc
     4141 ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg
     4201 agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg
     4261 atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc
     4321 acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg
     4381 aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga
     4441 gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact
     4501 ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga
     4561 caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc accgtgcgga
     4621 aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct
     4681 tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc agaaagaagg
     4741 actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg
     4801 tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg
     4861 ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca
     4921 agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg
     4981 agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg
     5041 aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc
     5101 tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaagcact
     5161 acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg
     5221 ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc atcagagagc
     5281 aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca
     5341 agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg
     5401 ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc
     5461 tgggaggcga caaaaggccg gcggccacga aaaaggccgg ccaggcaaaa aagaaaaagt
     5521 aagaattcct agagctcgct gatcagcctc gactgtgcct tctagttgcc agccatctgt
     5581 tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca ctgtcctttc
     5641 ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta ttctgggggg
     5701 tggggtgggg caggacagca agggggagga ttgggaagag aatagcaggc atgctgggga
     5761 gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg ctcgctcgct
     5821 cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg cggcctcagt
     5881 gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc tccttacgca
     5941 tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc
     6001 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     6061 ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     6121 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     6181 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     6241 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     6301 ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     6361 tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     6421 atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga tgccgcatag
     6481 ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     6541 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     6601 tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct atttttatag
     6661 gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg
     6721 cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga
     6781 caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat
     6841 ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca
     6901 gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc
     6961 gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca
     7021 atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg
     7081 caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca
     7141 gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata
     7201 accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag
     7261 ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg
     7321 gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca
     7381 acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     7441 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     7501 ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg tatcattgca
     7561 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     7621 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     7681 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt
     7741 taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa
     7801 cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga
     7861 gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg
     7921 gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc
     7981 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
     8041 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
     8101 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
     8161 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
     8221 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
     8281 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
     8341 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
     8401 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
     8461 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
