LOCUS       sequence-391667-        9223 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression construct encoding the SpCas9-D10A nickase.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9223)
  AUTHORS   Wang Q, Liu J, Janssen JM, Le Bouteiller M, Frock RL, Goncalves MAFV
  TITLE     Precise and broad scope genome editing based on high-specificity 
            Cas9 nickases.
  JOURNAL   Nucleic Acids Res. 2021 Jan 25;49(2):1173-1198. doi: 
            10.1093/nar/gkaa1236.
  PUBMED    33398349
REFERENCE   2  (bases 1 to 9223)
  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"; doi: "10"; journalName: 
            "Nucleic Acids Res"; date: "2021-01-25- 25"; volume: "49"; issue: 
            "2"; pages: "1173-1198"
COMMENT     Sequence Label: AB65_pCAG.Cas9-D10A.rBGpA
FEATURES             Location/Qualifiers
     source          1..9223
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        33..412
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        414..689
                     /label=chicken beta-actin promoter
     intron          690..1706
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1630..1652
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1714..1733
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1804..1813
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1810..1836,1837..1839,1840..5913)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             1810..5913
                     /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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5926..5946
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(6090..6109)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    6155..6210
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(6209..6228)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(6589..6606)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6589..6605)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6598..6620)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(6814..6833)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6933..6955
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6993..7011)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7079..7183
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7184..7252,7253..8044)
                     /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(7402..7421)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8215..8803
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8704..8723
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8957..8974
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    9091..9112
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(9127..9132,9133..9150,9151..9157)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9165..9181
                     /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     9170..9192
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     9189..9205
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9189..9205
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 gtttaaacat ttaaatctcg agccatggat tcgacattga ttattgacta gttattaata
       61 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact
      121 tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat
      181 gacgtatgtt cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta
      241 tttacggtaa actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc
      301 tattgacgtc aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg
      361 ggactttcct acttggcagt acatctacgt attagtcatc gctattacca tggtcgaggt
      421 gagccccacg ttctgcttca ctctccccat ctcccccccc tccccacccc caattttgta
      481 tttatttatt ttttaattat tttgtgcagc gatgggggcg gggggggggg gggsgcgcgc
      541 caggcggggc ggggcggggc gaggggcggg gcggggcgag gcggagaggt gcggcggcag
      601 ccaatcagag cggcgcgctc cgaaagtttc cttttatggc gaggcggcgg cggcggcggc
      661 cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc gcgctgcctt cgccccgtgc
      721 cccgctccgc cgccgcctcg cgccgcccgc cccggctctg actgaccgcg ttactcccac
      781 aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa ttagcgcttg gtttaatgac
      841 ggcttgtttc ttttctgtgg ctgcgtgaaa gccttgaggg gctccgggag ggccctttgt
      901 gcggggggag cggctcgggg ggtgcgtgcg tgtgtgtgtg cgtggggagc gccgcgtgcg
      961 gctccgcgct gcccggcggc tgtgagcgct gcgggcgcgg cgcggggctt tgtgcgctcc
     1021 gcagtgtgcg cgaggggagc gcggccgggg gcggtgcccc gcggtgcggg gggggctgcg
     1081 aggggaacaa aggctgcgtg cggggtgtgt gcgtgggggg gtgagcaggg ggtgtgggcg
     1141 cgtcggtcgg gctgcaaccc cccctgcacc cccctccccg agttgctgag cacggcccgg
     1201 cttcgggtgc ggggctccgt acggggcgtg gcgcggggct cgccgtgccg ggcggggggt
     1261 ggcggcaggt gggggtgccg ggcggggcgg ggccgcctcg ggccggggag ggctcggggg
     1321 aggggcgcgg cggcccccgg agcgccggcg gctgtcgagg cgcggcgagc cgcagccatt
     1381 gccttttatg gtaatcgtgc gagagggcgc agggacttcc tttgtcccaa atctgtgcgg
     1441 agccgaaatc tgggaggcgc cgccgcaccc cctctagcgg gcgcggggcg aagcggtgcg
     1501 gcgccggcag gaaggaaatg ggcggggagg gccttcgtgc gtcgccgcgc cgccgtcccc
     1561 ttctccctct ccagcctcgg ggctgtccgc ggggggacgg ctgccttcgg gggggacggg
     1621 gcagggcggg gttcggcttc tggcgtgtga ccggcggctc tagagcctct gctaaccatg
     1681 ttcatgcctt cttctttttc ctacagctcc tgggcaacgt gctggttatt gtgctgtctc
     1741 atcattttgg caaagaatta tcgcatgcct gcagagctct agagtctaga ggatcgaacc
     1801 cttgccacca tggacaagaa gtactccatt gggctcgcta tcggcacaaa cagcgtcggc
     1861 tgggccgtca ttacggacga gtacaaggtg ccgagcaaaa aattcaaagt tctgggcaat
     1921 accgatcgcc acagcataaa gaagaacctc attggcgccc tcctgttcga ctccggggag
     1981 acggccgaag ccacgcggct caaaagaaca gcacggcgca gatatacccg cagaaagaat
     2041 cggatctgct acctgcagga gatctttagt aatgagatgg ctaaggtgga tgactctttc
     2101 ttccataggc tggaggagtc ctttttggtg gaggaggata aaaagcacga gcgccaccca
     2161 atctttggca atatcgtgga cgaggtggcg taccatgaaa agtacccaac catatatcat
     2221 ctgaggaaga agcttgtaga cagtactgat aaggctgact tgcggttgat ctatctcgcg
     2281 ctggcgcata tgatcaaatt tcggggacac ttcctcatcg agggggacct gaacccagac
     2341 aacagcgatg tcgacaaact ctttatccaa ctggttcaga cttacaatca gcttttcgaa
     2401 gagaacccga tcaacgcatc cggagttgac gccaaagcaa tcctgagcgc taggctgtcc
     2461 aaatcccggc ggctcgaaaa cctcatcgca cagctccctg gggagaagaa gaacggcctg
     2521 tttggtaatc ttatcgccct gtcactcggg ctgaccccca actttaaatc taacttcgac
     2581 ctggccgaag atgccaagct tcaactgagc aaagacacct acgatgatga tctcgacaat
     2641 ctgctggccc agatcggcga ccagtacgca gacctttttt tggcggcaaa gaacctgtca
     2701 gacgccattc tgctgagtga tattctgcga gtgaacacgg agatcaccaa agctccgctg
     2761 agcgctagta tgatcaagcg ctatgatgag caccaccaag acttgacttt gctgaaggcc
     2821 cttgtcagac agcaactgcc tgagaagtac aaggaaattt tcttcgatca gtctaaaaat
     2881 ggctacgccg gatacattga cggcggagca agccaggagg aattttacaa atttattaag
     2941 cccatcttgg aaaaaatgga cggcaccgag gagctgctgg taaagcttaa cagagaagat
     3001 ctgttgcgca aacagcgcac tttcgacaat ggaagcatcc cccaccagat tcacctgggc
     3061 gaactgcacg ctatcctcag gcggcaagag gatttctacc cctttttgaa agataacagg
     3121 gaaaagattg agaaaatcct cacatttcgg ataccctact atgtaggccc cctcgcccgg
     3181 ggaaattcca gattcgcgtg gatgactcgc aaatcagaag agaccatcac tccctggaac
     3241 ttcgaggaag tcgtggataa gggggcctct gcccagtcct tcatcgaaag gatgactaac
     3301 tttgataaaa atctgcctaa cgaaaaggtg cttcctaaac actctctgct gtacgagtac
     3361 ttcacagttt ataacgagct caccaaggtc aaatacgtca cagaagggat gagaaagcca
     3421 gcattcctgt ctggagagca gaagaaagct atcgtggacc tcctcttcaa gacgaaccgg
     3481 aaagttaccg tgaaacagct caaagaagac tatttcaaaa agattgaatg tttcgactct
     3541 gttgaaatca gcggagtgga ggatcgcttc aacgcatccc tgggaacgta tcacgatctc
     3601 ctgaaaatca ttaaagacaa ggacttcctg gacaatgagg agaacgagga cattcttgag
     3661 gacattgtcc tcacccttac gttgtttgaa gatagggaga tgattgaaga acgcttgaaa
     3721 acttacgctc atctcttcga cgacaaagtc atgaaacagc tcaagaggcg ccgatataca
     3781 ggatgggggc ggctgtcaag aaaactgatc aatgggatcc gagacaagca gagtggaaag
     3841 acaatcctgg attttcttaa gtccgatgga tttgccaacc ggaacttcat gcagttgatc
     3901 catgatgact ctctcacctt taaggaggac atccagaaag cacaagtttc tggccagggg
     3961 gacagtcttc acgagcacat cgctaatctt gcaggtagcc cagctatcaa aaagggaata
     4021 ctgcagaccg ttaaggtcgt ggatgaactc gtcaaagtaa tgggaaggca taagcccgag
     4081 aatatcgtta tcgagatggc ccgagagaac caaactaccc agaagggaca gaagaacagt
     4141 agggaaagga tgaagaggat tgaagagggt ataaaagaac tggggtccca aatccttaag
     4201 gaacacccag ttgaaaacac ccagcttcag aatgagaagc tctacctgta ctacctgcag
     4261 aacggcaggg acatgtacgt ggatcaggaa ctggacatca atcggctctc cgactacgac
     4321 gtggatcata tcgtgcccca gtcttttctc aaagatgatt ctattgataa taaagtgttg
     4381 acaagatccg ataaaaatag agggaagagt gataacgtcc cctcagaaga agttgtcaag
     4441 aaaatgaaaa attattggcg gcagctgctg aacgccaaac tgatcacaca acggaagttc
     4501 gataatctga ctaaggctga acgaggtggc ctgtctgagt tggataaagc cggcttcatc
     4561 aaaaggcagc ttgttgagac acgccagatc accaagcacg tggcccaaat tctcgattca
     4621 cgcatgaaca ccaagtacga tgaaaatgac aaactgattc gagaggtgaa agttattact
     4681 ctgaagtcta agctggtctc agatttcaga aaggactttc agttttataa ggtgagagag
     4741 atcaacaatt accaccatgc gcatgatgcc tacctgaatg cagtggtagg cactgcactt
     4801 atcaaaaaat atcccaagct tgaatctgaa tttgtttacg gagactataa agtgtacgat
     4861 gttaggaaaa tgatcgcaaa gtctgagcag gaaataggca aggccaccgc taagtacttc
     4921 ttttacagca atattatgaa ttttttcaag accgagatta cactggccaa tggagagatt
     4981 cggaagcgac cacttatcga aacaaacgga gaaacaggag aaatcgtgtg ggacaagggt
     5041 agggatttcg cgacagtccg gaaggtcctg tccatgccgc aggtgaacat cgttaaaaag
     5101 accgaagtac agaccggagg cttctccaag gaaagtatcc tcccgaaaag gaacagcgac
     5161 aagctgatcg cacgcaaaaa agattgggac cccaagaaat acggcggatt cgattctcct
     5221 acagtcgctt acagtgtact ggttgtggcc aaagtggaga aagggaagtc taaaaaactc
     5281 aaaagcgtca aggaactgct gggcatcaca atcatggagc gatcaagctt cgaaaaaaac
     5341 cccatcgact ttctcgaggc gaaaggatat aaagaggtca aaaaagacct catcattaag
     5401 cttcccaagt actctctctt tgagcttgaa aacggccgga aacgaatgct cgctagtgcg
     5461 ggcgagctgc agaaaggtaa cgagctggca ctgccctcta aatacgttaa tttcttgtat
     5521 ctggccagcc actatgaaaa gctcaaaggg tctcccgaag ataatgagca gaagcagctg
     5581 ttcgtggaac aacacaaaca ctaccttgat gagatcatcg agcaaataag cgaattctcc
     5641 aaaagagtga tcctcgccga cgctaacctc gataaggtgc tttctgctta caataagcac
     5701 agggataagc ccatcaggga gcaggcagaa aacattatcc acttgtttac tctgaccaac
     5761 ttgggcgcgc ctgcagcctt caagtacttc gacaccacca tagacagaaa gcggtacacc
     5821 tctacaaagg aggtcctgga cgccacactg attcatcagt caattacggg gctctatgaa
     5881 acaagaatcg acctctctca gctcggtgga gacagcaggg ctgaccccaa gaagaagagg
     5941 aaggtgtgaa agggttcgat ccctaccggt tagtaatgag tttggccgca cttaagttac
     6001 gcgtggaaat tcactcctca ggtgcaggct gcctatcaga aggtggtggc tggtgtggcc
     6061 aatgccctgg ctcacaaata ccactgagat ctttttccct ctgccaaaaa ttatggggac
     6121 atcatgaagc cccttgagca tctgacttct ggctaataaa ggaaatttat tttcattgca
     6181 atagtgtgtt ggaatttttt gtgtctctca ctcggaagga catatgggag ggcaaatcat
     6241 ttaaaacatc agaatgagta tttggtttag agtttggcaa catatgccca tatgctggct
     6301 gccatgaaca aaggttggct ataaagaggt catcagtata tgaaacagcc ccctgctgtc
     6361 cattccttat tccatagaaa agccttgact tgaggttaga ttttttttat attttgtttt
     6421 gtgttatttt tttctttaac atccctaaaa ttttccttac atgttttact agccagattt
     6481 ttcctcctct cctgactact cccagtcata gctgtccctc ttctcttatg gagatccctc
     6541 gacctgcagc ccaagctgat cccgggattt aaatgtttaa acgaattcac tggccgtcgt
     6601 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
     6661 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
     6721 gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta cgcatctgtg
     6781 cggtatttca caccgcatat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt
     6841 aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc
     6901 ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc
     6961 accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt
     7021 taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg
     7081 cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca
     7141 ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt
     7201 ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga
     7261 aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga
     7321 actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat
     7381 gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca
     7441 agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt
     7501 cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac
     7561 catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct
     7621 aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga
     7681 gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac
     7741 aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat
     7801 agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg
     7861 ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc
     7921 actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc
     7981 aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg
     8041 gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta
     8101 atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg
     8161 tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga
     8221 tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt
     8281 ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag
     8341 agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa
     8401 ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag
     8461 tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca
     8521 gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac
     8581 cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa
     8641 ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc
     8701 agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg
     8761 tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc
     8821 ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc
     8881 ccctgattct gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag
     8941 ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa
     9001 accgcctctc cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga
     9061 ctggaaagcg ggcagtgagc gcaacgcaat taatgtgagt tagctcactc attaggcacc
     9121 ccaggcttta cactttatgc ttccggctcg tatgttgtgt ggaattgtga gcggataaca
     9181 atttcacaca ggaaacagct atgaccatga ttacgccaag ctt
//
