LOCUS       sequence-259494-        9673 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression vector for sgRNAs cloned into the BsaI sites and for 
            expression of Cas9 with 126aa MRN-recruiting domain from HSV-1 UL12 
            fused to N-terminus of Cas9, linked to mCherry via a T2A peptide.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9673)
  AUTHORS   Reuven N, Adler J, Broennimann K, Myers N, Shaul Y
  TITLE     Recruitment of DNA Repair MRN Complex by Intrinsically Disordered 
            Protein Domain Fused to Cas9 Improves Efficiency of CRISPR-Mediated 
            Genome Editing.
  JOURNAL   Biomolecules. 2019 Oct 8;9(10). pii: biom9100584. doi: 
            10.3390/biom9100584.
  PUBMED    31597252
REFERENCE   2  (bases 1 to 9673)
  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: 
            "Biomolecules."; date: "2019-10-8"; volume: "9(10). pii: 
            biom9100584. doi"; pages: " 10.3390/biom9100584"
COMMENT     Sequence Label: pU6-(BsaI)_CBh-UN-Cas9-T2A-mCherry
FEATURES             Location/Qualifiers
     source          1..9673
                     /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        270..345
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        442..727
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        729..1006
                     /label=chicken beta-actin promoter
     intron          1007..1235
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1254..1263
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1650..1715
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1722..1742
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1767..5867
                     /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             1767..5867
                     /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             5868..5915
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5919..5972
                     /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             5979..6689
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(6124..6142)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(6714..6731)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6720..6927
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6936..7076
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6936..7065
                     /label=AAV2 ITR
     rep_origin      7151..7606
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7238..7257)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7448..7469
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7623..7642)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7742..7764
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7802..7820)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7888..7992
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7993..8061,8062..8853)
                     /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(8211..8230)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9024..9612
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9513..9532
                     /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 gagacctgta caggtctctg ttttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tgttttagag
      361 ctagaaatag caagttaaaa taaggctagt ccgtttttag cgcgtgcgcc aattctgcag
      421 acaaatggct ctagaggtac ccgttacata acttacggta aatggcccgc ctggctgacc
      481 gcccaacgac ccccgcccat tgacgtcaat agtaacgcca atagggactt tccattgacg
      541 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
      601 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attgtgccca
      661 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
      721 taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc
      781 acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg
      841 gggggggggg cgcgcgccag gcggggcggg gcggggcgag gggcggggcg gggcgaggcg
      901 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
      961 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgac
     1021 gctgccttcg ccccgtgccc cgctccgccg ccgcctcgcg ccgcccgccc cggctctgac
     1081 tgaccgcgtt actcccacag gtgagcgggc gggacggccc ttctcctccg ggctgtaatt
     1141 agctgagcaa gaggtaaggg tttaagggat ggttggttgg tggggtatta atgtttaatt
     1201 acctggagca cctgcctgaa atcacttttt ttcaggttgg accggtgaag cttgccacca
     1261 tggagtccac gggaggccca gcatgtccgc cgggacgcac cgtgactaag cgttcctggg
     1321 ccctggccga ggacacccct cgtggccccg acagcccccc caagcgcccc cgccctaaca
     1381 gtcttccgct aacaaccacc ttccgtcccc tgcccccccc accccagacg acgtcagctg
     1441 tggacccgag ctcccattcg cccgttaacc ccccacgtga tcagcacgcc accgacaccg
     1501 cagacgaaaa gccccgggcc gcgtcgccgg cactttctga cgcctcaggg cctccgaccc
     1561 cagacattcc gctatctcct gggggcaccc acgcccgcga cccggacgcc gatcccgact
     1621 ccccggacct tgactctgga tccgtcatgg actataagga ccacgacgga gactacaagg
     1681 atcatgatat tgattacaaa gacgatgacg ataagatggc cccaaagaag aagcggaagg
     1741 tcggtatcca cggagtccca gcagccgaca agaagtacag catcggcctg gacatcggca
     1801 ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca
     1861 aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt
     1921 tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca
     1981 ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg
     2041 tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc
     2101 acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc
     2161 ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc
     2221 tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg
     2281 acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca
     2341 accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt
     2401 ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga
     2461 agaagaatgg cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca
     2521 agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg
     2581 acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg
     2641 ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca
     2701 ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga
     2761 ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg
     2821 accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct
     2881 acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc
     2941 tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc
     3001 agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc
     3061 tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg
     3121 gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca
     3181 tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg
     3241 agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc
     3301 tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg
     3361 gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt
     3421 tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg
     3481 agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca
     3541 cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg
     3601 aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg
     3661 aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc
     3721 ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca
     3781 agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact
     3841 tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg
     3901 tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca
     3961 ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc
     4021 ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg
     4081 gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca
     4141 gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc
     4201 tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc
     4261 tgtccgacta cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg
     4321 acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg
     4381 aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta
     4441 cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata
     4501 aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac
     4561 agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag
     4621 tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt
     4681 acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg
     4741 tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact
     4801 acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta
     4861 ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg
     4921 ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg
     4981 tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga
     5041 atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca
     5101 agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg
     5161 gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca
     5221 agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca
     5281 gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg
     5341 acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa
     5401 tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg
     5461 tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg
     5521 agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga
     5581 tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg
     5641 cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt
     5701 ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc
     5761 ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca
     5821 ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaaa aggccggcgg
     5881 ccacgaaaaa ggccggccag gcaaaaaaga aaaagcttga gggcagagga agtctgctaa
     5941 catgcggtga cgtggaggag aatcccggcc ctgctagcat ggtgagcaag ggcgaggagg
     6001 ataacatggc catcatcaag gagttcatgc gcttcaaggt gcacatggag ggctccgtga
     6061 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     6121 ccgccaagct gaaggtgacc aagggtggcc ccctgccctt cgcctgggac atcctgtccc
     6181 ctcagttcat gtacggctcc aaggcctacg tgaagcaccc cgccgacatc cccgactact
     6241 tgaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     6301 gcgtggtgac cgtgacccag gactcctccc tgcaggacgg cgagttcatc tacaaggtga
     6361 agctgcgcgg caccaacttc ccctcagacg gccccgtaat gcagaagaaa accatgggct
     6421 gggaggcctc ctccgagcgg atgtaccccg aggacggcgc cctgaagggc gagatcaagc
     6481 agaggctgaa gctgaaggac ggcggccact acgacgctga ggtcaagacc acctacaagg
     6541 ccaagaagcc cgtgcagctg cccggcgcct acaacgtcaa catcaagttg gacatcacct
     6601 cccacaacga ggactacacc atcgtggaac agtacgaacg cgccgagggc cgccactcca
     6661 ccggcggcat ggacgagctg tacaagtaag aattcctaga gctcgctgat cagcctcgac
     6721 tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct
     6781 ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct
     6841 gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg
     6901 ggaagagaat agcaggcatg ctggggagcg gccgcaggaa cccctagtga tggagttggc
     6961 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     7021 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg
     7081 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag
     7141 caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     7201 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     7261 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     7321 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     7381 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     7441 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc gggctattct
     7501 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     7561 caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc
     7621 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     7681 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     7741 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     7801 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     7861 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     7921 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     7981 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     8041 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     8101 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     8161 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     8221 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     8281 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     8341 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     8401 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     8461 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     8521 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     8581 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     8641 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     8701 gagcgtggaa gccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     8761 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     8821 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     8881 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     8941 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     9001 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     9061 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     9121 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg
     9181 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     9241 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     9301 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     9361 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     9421 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     9481 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     9541 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     9601 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     9661 tttgctcaca tgt
//
