LOCUS       sequence-157221-        8132 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9 nickase in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8132)
  AUTHORS   Seirin-Lee S, Osakada F, Takeda J, Tashiro S, Kobayashi R, Yamamoto 
            T, Ochiai H
  TITLE     Role of dynamic nuclear deformation on genomic architecture 
            reorganization.
  JOURNAL   PLoS Comput Biol. 2019 Sep 11;15(9):e1007289. doi: 
            10.1371/journal.pcbi.1007289. eCollection 2019 Sep.
  PUBMED    31509522
REFERENCE   2  (bases 1 to 8132)
  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: "PLoS Comput
            Biol."; date: "2019-09-11"; volume: "15(9):e1007289. doi"; pages: " 
            10.1371/journal.pcbi.1007289. eCollection 2019 Sep"
COMMENT     Sequence Label: pBSK-Delta-B-CBh-Cas9n-pA
FEATURES             Location/Qualifiers
     source          1..8132
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     enhancer        659..944
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        946..1223
                     /label=chicken beta-actin promoter
     intron          1224..1452
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             1473..1499
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1503..1523
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1533..1556,1557..1559,1560..5633)
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1533..5633
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5637..5657
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5694..5711)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5700..5907
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(5944..5964)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(5944..5962)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(5983..5999)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5983..5999)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5996..6018)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6007..6023
                     /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(6031..6037,6038..6055,6056..6061))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6076..6097
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6214..6231)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6385..6973)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6465..6484)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7144..7935,7936..8004))
                     /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     7767..7786
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8005..8109)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tgggtacccg
      661 ttacataact tacggtaaat ggcccgcctg gctgaccgcc caacgacccc cgcccattga
      721 cgtcaatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      781 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      841 tcaatgacgg taaatggccc gcctggcatt gtgcccagta catgacctta tgggactttc
      901 ctacttggca gtacatctac gtattagtca tcgctattac catggtcgag gtgagcccca
      961 cgttctgctt cactctcccc atctcccccc cctccccacc cccaattttg tatttattta
     1021 ttttttaatt attttgtgca gcgatggggg cggggggggg gggggggcgc gcgccaggcg
     1081 gggcggggcg gggcgagggg cggggcgggg cgaggcggag aggtgcggcg gcagccaatc
     1141 agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata
     1201 aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgacgct gccttcgccc cgtgccccgc
     1261 tccgccgccg cctcgcgccg cccgccccgg ctctgactga ccgcgttact cccacaggtg
     1321 agcgggcggg acggcccttc tcctccgggc tgtaattagc tgagcaagag gtaagggttt
     1381 aagggatggt tggttggtgg ggtattaatg tttaattacc tggagcacct gcctgaaatc
     1441 actttttttc aggttggacc ggtgccacca tgtacccata cgatgttcca gattacgctt
     1501 cgccgaagaa aaagcgcaag gtcgaagcgt ccgacaagaa gtacagcatc ggcctggcca
     1561 tcggcaccaa ctctgtgggc tgggccgtga tcaccgacga gtacaaggtg cccagcaaga
     1621 aattcaaggt gctgggcaac accgaccggc acagcatcaa gaagaacctg atcggagccc
     1681 tgctgttcga cagcggcgaa acagccgagg ccacccggct gaagagaacc gccagaagaa
     1741 gatacaccag acggaagaac cggatctgct atctgcaaga gatcttcagc aacgagatgg
     1801 ccaaggtgga cgacagcttc ttccacagac tggaagagtc cttcctggtg gaagaggata
     1861 agaagcacga gcggcacccc atcttcggca acatcgtgga cgaggtggcc taccacgaga
     1921 agtaccccac catctaccac ctgagaaaga aactggtgga cagcaccgac aaggccgacc
     1981 tgcggctgat ctatctggcc ctggcccaca tgatcaagtt ccggggccac ttcctgatcg
     2041 agggcgacct gaaccccgac aacagcgacg tggacaagct gttcatccag ctggtgcaga
     2101 cctacaacca gctgttcgag gaaaacccca tcaacgccag cggcgtggac gccaaggcca
     2161 tcctgtctgc cagactgagc aagagcagac ggctggaaaa tctgatcgcc cagctgcccg
     2221 gcgagaagaa gaatggcctg ttcggcaacc tgattgccct gagcctgggc ctgaccccca
     2281 acttcaagag caacttcgac ctggccgagg atgccaaact gcagctgagc aaggacacct
     2341 acgacgacga cctggacaac ctgctggccc agatcggcga ccagtacgcc gacctgtttc
     2401 tggccgccaa gaacctgtcc gacgccatcc tgctgagcga catcctgaga gtgaacaccg
     2461 agatcaccaa ggcccccctg agcgcctcta tgatcaagag atacgacgag caccaccagg
     2521 acctgaccct gctgaaagct ctcgtgcggc agcagctgcc tgagaagtac aaagagattt
     2581 tcttcgacca gagcaagaac ggctacgccg gctacattga cggcggagcc agccaggaag
     2641 agttctacaa gttcatcaag cccatcctgg aaaagatgga cggcaccgag gaactgctcg
     2701 tgaagctgaa cagagaggac ctgctgcgga agcagcggac cttcgacaac ggcagcatcc
     2761 cccaccagat ccacctggga gagctgcacg ccattctgcg gcggcaggaa gatttttacc
     2821 cattcctgaa ggacaaccgg gaaaagatcg agaagatcct gaccttccgc atcccctact
     2881 acgtgggccc tctggccagg ggaaacagca gattcgcctg gatgaccaga aagagcgagg
     2941 aaaccatcac cccctggaac ttcgaggaag tggtggacaa gggcgcttcc gcccagagct
     3001 tcatcgagcg gatgaccaac ttcgataaga acctgcccaa cgagaaggtg ctgcccaagc
     3061 acagcctgct gtacgagtac ttcaccgtgt ataacgagct gaccaaagtg aaatacgtga
     3121 ccgagggaat gagaaagccc gccttcctga gcggcgagca gaaaaaggcc atcgtggacc
     3181 tgctgttcaa gaccaaccgg aaagtgaccg tgaagcagct gaaagaggac tacttcaaga
     3241 aaatcgagtg cttcgactcc gtggaaatct ccggcgtgga agatcggttc aacgcctccc
     3301 tgggcacata ccacgatctg ctgaaaatta tcaaggacaa ggacttcctg gacaatgagg
     3361 aaaacgagga cattctggaa gatatcgtgc tgaccctgac actgtttgag gacagagaga
     3421 tgatcgagga acggctgaaa acctatgccc acctgttcga cgacaaagtg atgaagcagc
     3481 tgaagcggcg gagatacacc ggctggggca ggctgagccg gaagctgatc aacggcatcc
     3541 gggacaagca gtccggcaag acaatcctgg atttcctgaa gtccgacggc ttcgccaaca
     3601 gaaacttcat gcagctgatc cacgacgaca gcctgacctt taaagaggac atccagaaag
     3661 cccaggtgtc cggccagggc gatagcctgc acgagcacat tgccaatctg gccggcagcc
     3721 ccgccattaa gaagggcatc ctgcagacag tgaaggtggt ggacgagctc gtgaaagtga
     3781 tgggccggca caagcccgag aacatcgtga tcgaaatggc cagagagaac cagaccaccc
     3841 agaagggaca gaagaacagc cgcgagagaa tgaagcggat cgaagagggc atcaaagagc
     3901 tgggcagcca gatcctgaaa gaacaccccg tggaaaacac ccagctgcag aacgagaagc
     3961 tgtacctgta ctacctgcag aatgggcggg atatgtacgt ggaccaggaa ctggacatca
     4021 accggctgtc cgactacgat gtggaccata tcgtgcctca gagctttctg aaggacgact
     4081 ccatcgacaa caaggtgctg accagaagcg acaagaaccg gggcaagagc gacaacgtgc
     4141 cctccgaaga ggtcgtgaag aagatgaaga actactggcg gcagctgctg aacgccaagc
     4201 tgattaccca gagaaagttc gacaatctga ccaaggccga gagaggcggc ctgagcgaac
     4261 tggataaggc cggcttcatc aagagacagc tggtggaaac ccggcagatc acaaagcacg
     4321 tggcacagat cctggactcc cggatgaaca ctaagtacga cgagaatgac aagctgatcc
     4381 gggaagtgaa agtgatcacc ctgaagtcca agctggtgtc cgatttccgg aaggatttcc
     4441 agttttacaa agtgcgcgag atcaacaact accaccacgc ccacgacgcc tacctgaacg
     4501 ccgtcgtggg aaccgccctg atcaaaaagt accctaagct ggaaagcgag ttcgtgtacg
     4561 gcgactacaa ggtgtacgac gtgcggaaga tgatcgccaa gagcgagcag gaaatcggca
     4621 aggctaccgc caagtacttc ttctacagca acatcatgaa ctttttcaag accgagatta
     4681 ccctggccaa cggcgagatc cggaagcggc ctctgatcga gacaaacggc gaaaccgggg
     4741 agatcgtgtg ggataagggc cgggattttg ccaccgtgcg gaaagtgctg agcatgcccc
     4801 aagtgaatat cgtgaaaaag accgaggtgc agacaggcgg cttcagcaaa gagtctatcc
     4861 tgcccaagag gaacagcgat aagctgatcg ccagaaagaa ggactgggac cctaagaagt
     4921 acggcggctt cgacagcccc accgtggcct attctgtgct ggtggtggcc aaagtggaaa
     4981 agggcaagtc caagaaactg aagagtgtga aagagctgct ggggatcacc atcatggaaa
     5041 gaagcagctt cgagaagaat cccatcgact ttctggaagc caagggctac aaagaagtga
     5101 aaaaggacct gatcatcaag ctgcctaagt actccctgtt cgagctggaa aacggccgga
     5161 agagaatgct ggcctctgcc ggcgaactgc agaagggaaa cgaactggcc ctgccctcca
     5221 aatatgtgaa cttcctgtac ctggccagcc actatgagaa gctgaagggc tcccccgagg
     5281 ataatgagca gaaacagctg tttgtggaac agcacaagca ctacctggac gagatcatcg
     5341 agcagatcag cgagttctcc aagagagtga tcctggccga cgctaatctg gacaaagtgc
     5401 tgtccgccta caacaagcac cgggataagc ccatcagaga gcaggccgag aatatcatcc
     5461 acctgtttac cctgaccaat ctgggagccc ctgccgcctt caagtacttt gacaccacca
     5521 tcgaccggaa gaggtacacc agcaccaaag aggtgctgga cgccaccctg atccaccaga
     5581 gcatcaccgg cctgtacgag acacggatcg acctgtctca gctgggaggc gacagcccca
     5641 agaagaagag aaaggtggag gccagctaag aattcctaga gctcgctgat cagcctcgac
     5701 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     5761 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     5821 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     5881 ggaagagaat agcaggcatg ctggggagcg gccgccaccg cggtggagct ccagcttttg
     5941 ttccctttag tgagggttaa ttgcgcgctt ggcgtaatca tggtcatagc tgtttcctgt
     6001 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     6061 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     6121 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     6181 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     6241 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     6301 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     6361 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     6421 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     6481 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     6541 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     6601 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     6661 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     6721 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     6781 tacagagttc ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat
     6841 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     6901 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     6961 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     7021 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     7081 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     7141 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     7201 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     7261 ccccagtgct gcaatgatac cgcggctgcc acgctcaccg gctccagatt tatcagcaat
     7321 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     7381 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     7441 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     7501 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     7561 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     7621 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     7681 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     7741 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     7801 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     7861 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     7921 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     7981 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     8041 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     8101 ggttccgcgc acatttcccc gaaaagtgcc ac
//
