LOCUS       sequence-315474-       10285 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SpCas9 and two sgRNA targeting the lenti-CDDR reporter at 
            two distal sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10285)
  AUTHORS   Eki R, She J, Parlak M, Benamar M, Du KP, Kumar P, Abbas T
  TITLE     A robust CRISPR-Cas9-based fluorescent reporter assay for the 
            detection and quantification of DNA double-strand break repair.
  JOURNAL   Nucleic Acids Res. 2020 Oct 17. pii: 5929239. doi: 
            10.1093/nar/gkaa897.
  PUBMED    33068408
REFERENCE   2  (bases 1 to 10285)
  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: "Nucleic 
            Acids Res. 2020 Oct 17. pii: 5929239. doi: 10.1093/nar/gkaa897."
COMMENT     Sequence Label: Cas9-sgRNA-A+B
FEATURES             Location/Qualifiers
     source          1..10285
                     /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        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..1234
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1246..1255
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1255..1320
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1327..1347
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1372..5472
                     /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             1372..5472
                     /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             5473..5520
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5548..5565)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5554..5761
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        5792..6040
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        6061..6136
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     protein_bind    6258..6291
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        6352..6851
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(6387..6410)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     6784..6803
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     CDS             6891..7490
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(6891..6910)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     rep_origin      7763..8218
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7850..7869)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     8060..8081
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(8235..8254)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8354..8376
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(8414..8432)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8500..8604
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8605..8673,8674..9465)
                     /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(8823..8842)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9636..10224
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     10125..10144
                     /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 gacggcggcg tgctgcagtg 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 cgcgcgccgg gggggggggg gggggggggg gggggggggg gggsgrggcg
      901 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
      961 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg
     1021 ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact
     1081 gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta
     1141 gctgagcaag aggtaagggt ttaagggatg gttggttggt ggggtattaa tgtttaatta
     1201 cctggagcac ctgcctgaaa tcactttttt tcaggttgga ccggtgccac catggactat
     1261 aaggaccacg acggagacta caaggatcat gatattgatt acaaagacga tgacgataag
     1321 atggccccaa agaagaagcg gaaggtcggt atccacggag tcccagcagc cgacaagaag
     1381 tacagcatcg gcctggacat cggcaccaac tctgtgggct gggccgtgat caccgacgag
     1441 tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca ccgaccggca cagcatcaag
     1501 aagaacctga tcggagccct gctgttcgac agcggcgaaa cagccgaggc cacccggctg
     1561 aagagaaccg ccagaagaag atacaccaga cggaagaacc ggatctgcta tctgcaagag
     1621 atcttcagca acgagatggc caaggtggac gacagcttct tccacagact ggaagagtcc
     1681 ttcctggtgg aagaggataa gaagcacgag cggcacccca tcttcggcaa catcgtggac
     1741 gaggtggcct accacgagaa gtaccccacc atctaccacc tgagaaagaa actggtggac
     1801 agcaccgaca aggccgacct gcggctgatc tatctggccc tggcccacat gatcaagttc
     1861 cggggccact tcctgatcga gggcgacctg aaccccgaca acagcgacgt ggacaagctg
     1921 ttcatccagc tggtgcagac ctacaaccag ctgttcgagg aaaaccccat caacgccagc
     1981 ggcgtggacg ccaaggccat cctgtctgcc agactgagca agagcagacg gctggaaaat
     2041 ctgatcgccc agctgcccgg cgagaagaag aatggcctgt tcggaaacct gattgccctg
     2101 agcctgggcc tgacccccaa cttcaagagc aacttcgacc tggccgagga tgccaaactg
     2161 cagctgagca aggacaccta cgacgacgac ctggacaacc tgctggccca gatcggcgac
     2221 cagtacgccg acctgtttct ggccgccaag aacctgtccg acgccatcct gctgagcgac
     2281 atcctgagag tgaacaccga gatcaccaag gcccccctga gcgcctctat gatcaagaga
     2341 tacgacgagc accaccagga cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct
     2401 gagaagtaca aagagatttt cttcgaccag agcaagaacg gctacgccgg ctacattgac
     2461 ggcggagcca gccaggaaga gttctacaag ttcatcaagc ccatcctgga aaagatggac
     2521 ggcaccgagg aactgctcgt gaagctgaac agagaggacc tgctgcggaa gcagcggacc
     2581 ttcgacaacg gcagcatccc ccaccagatc cacctgggag agctgcacgc cattctgcgg
     2641 cggcaggaag atttttaccc attcctgaag gacaaccggg aaaagatcga gaagatcctg
     2701 accttccgca tcccctacta cgtgggccct ctggccaggg gaaacagcag attcgcctgg
     2761 atgaccagaa agagcgagga aaccatcacc ccctggaact tcgaggaagt ggtggacaag
     2821 ggcgcttccg cccagagctt catcgagcgg atgaccaact tcgataagaa cctgcccaac
     2881 gagaaggtgc tgcccaagca cagcctgctg tacgagtact tcaccgtgta taacgagctg
     2941 accaaagtga aatacgtgac cgagggaatg agaaagcccg ccttcctgag cggcgagcag
     3001 aaaaaggcca tcgtggacct gctgttcaag accaaccgga aagtgaccgt gaagcagctg
     3061 aaagaggact acttcaagaa aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa
     3121 gatcggttca acgcctccct gggcacatac cacgatctgc tgaaaattat caaggacaag
     3181 gacttcctgg acaatgagga aaacgaggac attctggaag atatcgtgct gaccctgaca
     3241 ctgtttgagg acagagagat gatcgaggaa cggctgaaaa cctatgccca cctgttcgac
     3301 gacaaagtga tgaagcagct gaagcggcgg agatacaccg gctggggcag gctgagccgg
     3361 aagctgatca acggcatccg ggacaagcag tccggcaaga caatcctgga tttcctgaag
     3421 tccgacggct tcgccaacag aaacttcatg cagctgatcc acgacgacag cctgaccttt
     3481 aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg atagcctgca cgagcacatt
     3541 gccaatctgg ccggcagccc cgccattaag aagggcatcc tgcagacagt gaaggtggtg
     3601 gacgagctcg tgaaagtgat gggccggcac aagcccgaga acatcgtgat cgaaatggcc
     3661 agagagaacc agaccaccca gaagggacag aagaacagcc gcgagagaat gaagcggatc
     3721 gaagagggca tcaaagagct gggcagccag atcctgaaag aacaccccgt ggaaaacacc
     3781 cagctgcaga acgagaagct gtacctgtac tacctgcaga atgggcggga tatgtacgtg
     3841 gaccaggaac tggacatcaa ccggctgtcc gactacgatg tggaccatat cgtgcctcag
     3901 agctttctga aggacgactc catcgacaac aaggtgctga ccagaagcga caagaaccgg
     3961 ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg
     4021 cagctgctga acgccaagct gattacccag agaaagttcg acaatctgac caaggccgag
     4081 agaggcggcc tgagcgaact ggataaggcc ggcttcatca agagacagct ggtggaaacc
     4141 cggcagatca caaagcacgt ggcacagatc ctggactccc ggatgaacac taagtacgac
     4201 gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc
     4261 gatttccgga aggatttcca gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc
     4321 cacgacgcct acctgaacgc cgtcgtggga accgccctga tcaaaaagta ccctaagctg
     4381 gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg tgcggaagat gatcgccaag
     4441 agcgagcagg aaatcggcaa ggctaccgcc aagtacttct tctacagcaa catcatgaac
     4501 tttttcaaga ccgagattac cctggccaac ggcgagatcc ggaagcggcc tctgatcgag
     4561 acaaacggcg aaaccgggga gatcgtgtgg gataagggcc gggattttgc caccgtgcgg
     4621 aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc
     4681 ttcagcaaag agtctatcct gcccaagagg aacagcgata agctgatcgc cagaaagaag
     4741 gactgggacc ctaagaagta cggcggcttc gacagcccca ccgtggccta ttctgtgctg
     4801 gtggtggcca aagtggaaaa gggcaagtcc aagaaactga agagtgtgaa agagctgctg
     4861 gggatcacca tcatggaaag aagcagcttc gagaagaatc ccatcgactt tctggaagcc
     4921 aagggctaca aagaagtgaa aaaggacctg atcatcaagc tgcctaagta ctccctgttc
     4981 gagctggaaa acggccggaa gagaatgctg gcctctgccg gcgaactgca gaagggaaac
     5041 gaactggccc tgccctccaa atatgtgaac ttcctgtacc tggccagcca ctatgagaag
     5101 ctgaagggct cccccgagga taatgagcag aaacagctgt ttgtggaaca gcacaagcac
     5161 tacctggacg agatcatcga gcagatcagc gagttctcca agagagtgat cctggccgac
     5221 gctaatctgg acaaagtgct gtccgcctac aacaagcacc gggataagcc catcagagag
     5281 caggccgaga atatcatcca cctgtttacc ctgaccaatc tgggagcccc tgccgccttc
     5341 aagtactttg acaccaccat cgaccggaag aggtacacca gcaccaaaga ggtgctggac
     5401 gccaccctga tccaccagag catcaccggc ctgtacgaga cacggatcga cctgtctcag
     5461 ctgggaggcg acaaaaggcc ggcggccacg aaaaaggccg gccaggcaaa aaagaaaaag
     5521 taagaattcc tagagctcgc tgatcagcct cgactgtgcc ttctagttgc cagccatctg
     5581 ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt
     5641 cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg
     5701 gtggggtggg gcaggacagc aagggggagg attgggaaga gaatagcagg catgctgggg
     5761 agcggccgct gctggccttt tgctcacatg tgagggccta tttcccatga ttccttcata
     5821 tttgcatata cgatacaagg ctgttagaga gataattgga attaatttga ctgtaaacac
     5881 aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt agtttgcagt
     5941 tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa agtatttcga
     6001 tttcttggct ttatatatct tgtggaaagg acgaaacacc gtagctgaag ctcacgttca
     6061 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
     6121 ggcaccgagt cggtgctttt ttgttttaga gctagaaata gcaagttaaa ataaggctag
     6181 tccgttttta gcgcgtgcgc caattctgca gacaaatgta caagtaagaa ttccgatcat
     6241 attcaataac ccttaatata acttcgtata atgtatgcta tacgaagtta ttaggtctga
     6301 agaggagttt acgtccagcc aagcttagga tctcgacctc gaaattctac cgggtagggg
     6361 aggcgctttt cccaaggcag tctggagcat gcgctttagc agccccgctg ggcacttggc
     6421 gctacacaag tggcctctgg cctcgcacac attccacatc caccggtagg cgccaaccgg
     6481 ctccgttctt tggtggcccc ttcgcgccac cttctactcc tcccctagtc aggaagttcc
     6541 cccccgcccc gcagctcgcg tcgtgcagga cgtgacaaat ggaagtagca cgtctcacta
     6601 gtctcgtgca gatggacagc accgctgagc aatggaagcg ggtaggcctt tggggcagcg
     6661 gccaatagca gctttgctcc ttcgctttct gggctcagag gctgggaagg ggtgggtccg
     6721 ggggcgggct caggggcggg ctcaggggcg gggcgggcgc ccgaaggtcc tccggaggcc
     6781 cggcattctg cacgcttcaa aagcgcacgt ctgccgcgct gttctcctct tcctcatctc
     6841 cgggcctttc gacctgcatc catctagatc tcgagcagct gaagcttacc atgaccgagt
     6901 acaagcccac ggtgcgcctc gccacccgcg acgacgtccc cagggccgta cgcaccctcg
     6961 ccgccgcgtt cgccgactac cccgccacgc gccacaccgt cgatccggac cgccacatcg
     7021 agcgggtcac cgagctgcaa gaactcttcc tcacgcgcgt cgggctcgac atcggcaagg
     7081 tgtgggtcgc ggacgacggc gccgcggtgg cggtctggac cacgccggag agcgtcgaag
     7141 cgggggcggt gttcgccgag atcggcccgc gcatggccga gttgagcggt tcccggctgg
     7201 ccgcgcagca acagatggaa ggcctcctgg cgccgcaccg gcccaaggag cccgcgtggt
     7261 tcctggccac cgtcggcgtc tcgcccgacc accagggcaa gggtctgggc agcgccgtcg
     7321 tgctccccgg agtggaggcg gccgagcgcg ccggggtgcc cgccttcctg gagacctccg
     7381 cgccccacaa cctccccttc tacgagcggc tcggcttcac cgtcaccgcc gacgtcgagg
     7441 tgcccgaagg accgcgcacc tggtgcatga cccgcaagcc cggtgcctga cgcccgcccc
     7501 acgacccgca gcgcccgacc gaaaggagcg cacgacccca tgcatcgatg atatcagatc
     7561 cccgggatgc agaaattgat gatctattaa acaataaaga tgtccactaa aatggaagtt
     7621 tttcctgtca tactttgtta agaagggtga gaacagagta cctacatttt gaatggaagg
     7681 attggagcgg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     7741 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     7801 gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct
     7861 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     7921 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     7981 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     8041 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc
     8101 tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat
     8161 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     8221 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     8281 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     8341 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     8401 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     8461 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     8521 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     8581 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     8641 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     8701 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     8761 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     8821 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     8881 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     8941 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     9001 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     9061 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     9121 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     9181 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     9241 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     9301 tctggagccg gtgagcgtgg aagccgcggt atcattgcag cactggggcc agatggtaag
     9361 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     9421 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     9481 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     9541 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     9601 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     9661 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     9721 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     9781 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     9841 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     9901 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     9961 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
    10021 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
    10081 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
    10141 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
    10201 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
    10261 ttttgctggc cttttgctca catgt
//
