LOCUS       sequence-379152-        9723 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  All-in-one plasmid containing both guides for cutting BFP region in 
            DSB Spectrum V1.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9723)
  AUTHORS   van de Kooij B, Kruswick A, van Attikum H, Yaffe MB
  TITLE     Multi-pathway DNA-repair reporters reveal competition between 
            end-joining, single-strand annealing and homologous recombination at
            Cas9-induced DNA double-strand breaks.
  JOURNAL   Nat Commun. 2022 Sep 8;13(1):5295. doi: 10.1038/s41467-022-32743-w.
  PUBMED    36075911
REFERENCE   2  (bases 1 to 9723)
  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: "Nat 
            Commun."; date: "2022-09-8"; volume: "13(1):5295. doi"; pages: " 
            10.1038/s41467-022-32743-w"
COMMENT     Sequence Label: pSpCas9_T2A_mCherry sg1-sg2
FEATURES             Location/Qualifiers
     source          1..9723
                     /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)"
     misc_RNA        271..346
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        436..684
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        706..781
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        878..1163
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        1165..1442
                     /label=chicken beta-actin promoter
     intron          1443..1670
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1682..1691
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1691..1756
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1763..1783
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1808..5908
                     /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             1808..5908
                     /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             5909..5956
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5972..6025
                     /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             6026..6736
                     /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(6171..6189)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(6764..6781)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6770..6977
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6986..7126
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6986..7115
                     /label=AAV2 ITR
     rep_origin      7201..7656
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7288..7307)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7498..7519
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7673..7692)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7792..7814
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7852..7870)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7938..8042
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8043..8111,8112..8903)
                     /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(8261..8280)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9074..9662
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9563..9582
                     /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 accaccctct cccacggcta gttttagagc tagaaatagc aagttaaaat
      301 aaggctagtc cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgttttaga
      361 gctagaaata gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc caattctgca
      421 gacaaatggc tctaggaggg cctatttccc atgattcctt catatttgca tatacgatac
      481 aaggctgtta gagagataat tggaattaat ttgactgtaa acacaaagat attagtacaa
      541 aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt
      601 aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat
      661 atcttgtgga aaggacgaaa caccggggca aaacactgga ccaatgtttt agagctagaa
      721 atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg
      781 cttttttgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt ttttagcgcg
      841 tgcgccaatt ctgcagacaa atggctctag aggtacccgt tacataactt acggtaaatg
      901 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaatagta acgccaatag
      961 ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac
     1021 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
     1081 cctggcattg tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
     1141 tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca
     1201 tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag
     1261 cgatgggggc gggggggggg ggggggcgcg cgccaggcgg ggcggggcgg ggcgaggggc
     1321 ggggcggggc gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt
     1381 ttccttttat ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg
     1441 cgggagtcgc tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc
     1501 cgccccggct ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc
     1561 ctccgggctg taattagctg agcaagaggt aagggtttaa gggatggttg gttggtgggg
     1621 tattaatgtt taattacctg gagcacctgc ctgaaatcac tttttttcag gttggaccgg
     1681 tgccaccatg gactataagg accacgacgg agactacaag gatcatgata ttgattacaa
     1741 agacgatgac gataagatgg ccccaaagaa gaagcggaag gtcggtatcc acggagtccc
     1801 agcagccgac aagaagtaca gcatcggcct ggacatcggc accaactctg tgggctgggc
     1861 cgtgatcacc gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga
     1921 ccggcacagc atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc
     1981 cgaggccacc cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat
     2041 ctgctatctg caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca
     2101 cagactggaa gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt
     2161 cggcaacatc gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag
     2221 aaagaaactg gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc
     2281 ccacatgatc aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag
     2341 cgacgtggac aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa
     2401 ccccatcaac gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag
     2461 cagacggctg gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg
     2521 aaacctgatt gccctgagcc tgggcctgac ccccaacttc aagagcaact tcgacctggc
     2581 cgaggatgcc aaactgcagc tgagcaagga cacctacgac gacgacctgg acaacctgct
     2641 ggcccagatc ggcgaccagt acgccgacct gtttctggcc gccaagaacc tgtccgacgc
     2701 catcctgctg agcgacatcc tgagagtgaa caccgagatc accaaggccc ccctgagcgc
     2761 ctctatgatc aagagatacg acgagcacca ccaggacctg accctgctga aagctctcgt
     2821 gcggcagcag ctgcctgaga agtacaaaga gattttcttc gaccagagca agaacggcta
     2881 cgccggctac attgacggcg gagccagcca ggaagagttc tacaagttca tcaagcccat
     2941 cctggaaaag atggacggca ccgaggaact gctcgtgaag ctgaacagag aggacctgct
     3001 gcggaagcag cggaccttcg acaacggcag catcccccac cagatccacc tgggagagct
     3061 gcacgccatt ctgcggcggc aggaagattt ttacccattc ctgaaggaca accgggaaaa
     3121 gatcgagaag atcctgacct tccgcatccc ctactacgtg ggccctctgg ccaggggaaa
     3181 cagcagattc gcctggatga ccagaaagag cgaggaaacc atcaccccct ggaacttcga
     3241 ggaagtggtg gacaagggcg cttccgccca gagcttcatc gagcggatga ccaacttcga
     3301 taagaacctg cccaacgaga aggtgctgcc caagcacagc ctgctgtacg agtacttcac
     3361 cgtgtataac gagctgacca aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt
     3421 cctgagcggc gagcagaaaa aggccatcgt ggacctgctg ttcaagacca accggaaagt
     3481 gaccgtgaag cagctgaaag aggactactt caagaaaatc gagtgcttcg actccgtgga
     3541 aatctccggc gtggaagatc ggttcaacgc ctccctgggc acataccacg atctgctgaa
     3601 aattatcaag gacaaggact tcctggacaa tgaggaaaac gaggacattc tggaagatat
     3661 cgtgctgacc ctgacactgt ttgaggacag agagatgatc gaggaacggc tgaaaaccta
     3721 tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag cggcggagat acaccggctg
     3781 gggcaggctg agccggaagc tgatcaacgg catccgggac aagcagtccg gcaagacaat
     3841 cctggatttc ctgaagtccg acggcttcgc caacagaaac ttcatgcagc tgatccacga
     3901 cgacagcctg acctttaaag aggacatcca gaaagcccag gtgtccggcc agggcgatag
     3961 cctgcacgag cacattgcca atctggccgg cagccccgcc attaagaagg gcatcctgca
     4021 gacagtgaag gtggtggacg agctcgtgaa agtgatgggc cggcacaagc ccgagaacat
     4081 cgtgatcgaa atggccagag agaaccagac cacccagaag ggacagaaga acagccgcga
     4141 gagaatgaag cggatcgaag agggcatcaa agagctgggc agccagatcc tgaaagaaca
     4201 ccccgtggaa aacacccagc tgcagaacga gaagctgtac ctgtactacc tgcagaatgg
     4261 gcgggatatg tacgtggacc aggaactgga catcaaccgg ctgtccgact acgatgtgga
     4321 ccatatcgtg cctcagagct ttctgaagga cgactccatc gacaacaagg tgctgaccag
     4381 aagcgacaag aaccggggca agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat
     4441 gaagaactac tggcggcagc tgctgaacgc caagctgatt acccagagaa agttcgacaa
     4501 tctgaccaag gccgagagag gcggcctgag cgaactggat aaggccggct tcatcaagag
     4561 acagctggtg gaaacccggc agatcacaaa gcacgtggca cagatcctgg actcccggat
     4621 gaacactaag tacgacgaga atgacaagct gatccgggaa gtgaaagtga tcaccctgaa
     4681 gtccaagctg gtgtccgatt tccggaagga tttccagttt tacaaagtgc gcgagatcaa
     4741 caactaccac cacgcccacg acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa
     4801 aaagtaccct aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg
     4861 gaagatgatc gccaagagcg agcaggaaat cggcaaggct accgccaagt acttcttcta
     4921 cagcaacatc atgaactttt tcaagaccga gattaccctg gccaacggcg agatccggaa
     4981 gcggcctctg atcgagacaa acggcgaaac cggggagatc gtgtgggata agggccggga
     5041 ttttgccacc gtgcggaaag tgctgagcat gccccaagtg aatatcgtga aaaagaccga
     5101 ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc aagaggaaca gcgataagct
     5161 gatcgccaga aagaaggact gggaccctaa gaagtacggc ggcttcgaca gccccaccgt
     5221 ggcctattct gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga aactgaagag
     5281 tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc agcttcgaga agaatcccat
     5341 cgactttctg gaagccaagg gctacaaaga agtgaaaaag gacctgatca tcaagctgcc
     5401 taagtactcc ctgttcgagc tggaaaacgg ccggaagaga atgctggcct ctgccggcga
     5461 actgcagaag ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc tgtacctggc
     5521 cagccactat gagaagctga agggctcccc cgaggataat gagcagaaac agctgtttgt
     5581 ggaacagcac aagcactacc tggacgagat catcgagcag atcagcgagt tctccaagag
     5641 agtgatcctg gccgacgcta atctggacaa agtgctgtcc gcctacaaca agcaccggga
     5701 taagcccatc agagagcagg ccgagaatat catccacctg tttaccctga ccaatctggg
     5761 agcccctgcc gccttcaagt actttgacac caccatcgac cggaagaggt acaccagcac
     5821 caaagaggtg ctggacgcca ccctgatcca ccagagcatc accggcctgt acgagacacg
     5881 gatcgacctg tctcagctgg gaggcgacaa aaggccggcg gccacgaaaa aggccggcca
     5941 ggcaaaaaag aaaaaggaat tcggcagtgg agagggcaga ggaagtctgc taacatgcgg
     6001 tgacgtcgag gagaatcctg gcccaatggt gagcaagggc gaggaggata acatggccat
     6061 catcaaggag ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt
     6121 cgagatcgag ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa
     6181 ggtgaccaag ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta
     6241 cggctccaag gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt
     6301 ccccgagggc ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt
     6361 gacccaggac tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac
     6421 caacttcccc tccgacggcc ccgtaatgca gaagaaaacc atgggctggg aggcctcctc
     6481 cgagcggatg taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct
     6541 gaaggacggc ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt
     6601 gcagctgccc ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga
     6661 ctacaccatc gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga
     6721 cgagctgtac aagtgagaat tctaactaga gctcgctgat cagcctcgac tgtgccttct
     6781 agttgccagc catctgttgt ttgcccctcc cccgtgcctt ccttgaccct ggaaggtgcc
     6841 actcccactg tcctttccta ataaaatgag gaaattgcat cgcattgtct gagtaggtgt
     6901 cattctattc tggggggtgg ggtggggcag gacagcaagg gggaggattg ggaagagaat
     6961 agcaggcatg ctggggagcg gccgcaggaa cccctagtga tggagttggc cactccctct
     7021 ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg cccgggcttt
     7081 gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg cctgatgcgg
     7141 tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag caaccatagt
     7201 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     7261 ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     7321 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     7381 gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca cgtagtgggc
     7441 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     7501 gactcttgtt ccaaactgga acaacactca actctatctc gggctattct tttgatttat
     7561 aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa caaaaattta
     7621 acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc agtacaatct
     7681 gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct
     7741 gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct
     7801 gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga
     7861 tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca
     7921 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     7981 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     8041 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     8101 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     8161 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     8221 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     8281 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     8341 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     8401 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     8461 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     8521 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     8581 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     8641 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     8701 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtggaa
     8761 gccgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     8821 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     8881 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     8941 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     9001 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     9061 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     9121 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     9181 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     9241 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     9301 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     9361 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     9421 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     9481 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     9541 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     9601 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     9661 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     9721 tgt
//
