LOCUS       sequence-371632-       11565 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PiggyBac compatible plasmid expressing spCas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11565)
  AUTHORS   Nghi Dang , Alissa Lance-Byrne, Angela Tung, Xiaoge Guo, Ryan J 
            Cecchi, Joanna Buchthal, Alejandro Chavez, Nan Cher Yeo
  TITLE     Generation and application of a versatile CRISPR toolkit for 
            mammalian cell engineering
  JOURNAL   Synthetic Biology, Volume 6, Issue 1, 2021
REFERENCE   2  (bases 1 to 11565)
  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: "Synthetic 
            Biology, Volume 6, Issue 1, 2021"
COMMENT     Sequence Label: NAX_CAGGS_CAS9_mcherry
FEATURES             Location/Qualifiers
     source          1..11565
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     537..559
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     621..640
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      709..718
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             715..4818
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             715..4818
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             715..4818
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             4831..4851
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(4906..4925)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    4950..4998
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        5230..5441
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      5451..5460
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(5575..5593)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     5843..5862
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    6144..6265
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6181..6200)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6235..6254
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   7028..7062
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7553..7569)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7553..7569)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7566..7588)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7577..7593
                     /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(7601..7607,7608..7625,7626..7631))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7646..7667
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7784..7801)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7955..8543)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8035..8054)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8714..9505,9506..9574))
                     /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     9337..9356
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9575..9679)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(9705..10160)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9842..9863)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     10054..10073
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10287..10309
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     10301..10318
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     10302..10318
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   10448..10510
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        10931..11310
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg
       61 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg
      121 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg
      181 ggggsggggg ggggsggggc cgccksgggc sggggrgggc tcgggggagg ggcgcggcgg
      241 cccccggagc gccggcggct gtcgaggcgc ggcgagccgc agccattgcc ttttatggta
      301 atcgtgcgag agggcgcagg gacttccttt gtcccaaatc tgtgcggagc cgaaatctgg
      361 gaggcgccgc cgcaccccct ctagcgggcg cggggcgaag cggtgcggcg ccggcaggaa
      421 ggaaatgggc ggggagggcc ttcgtgcgtc gccgcgccgc cgtccccttc tccctctcca
      481 gcctcggggc tgtccgcggg gggacggctg ccttcggggg ggacggggca gggcggggtt
      541 cggcttctgg cgtgtgaccg gcggctctag agcctctgct aaccatgttc atgccttctt
      601 ctttttccta cagctcctgg gcaacgtgct ggttattgtg ctgtctcatc attttggcaa
      661 agaatctaga taccctcgta aaggtctaga gctagcgaat tcgaatttgc caccatggac
      721 aagaagtact ccattgggct cgatatcggc acaaacagcg tcggctgggc cgtcattacg
      781 gacgagtaca aggtgccgag caaaaaattc aaagttctgg gcaataccga tcgccacagc
      841 ataaagaaga acctcattgg cgccctcctg ttcgactccg gggagacggc cgaagccacg
      901 cggctcaaaa gaacagcacg gcgcagatat acccgcagaa agaatcggat ctgctacctg
      961 caggagatct ttagtaatga gatggctaag gtggatgact ctttcttcca taggctggag
     1021 gagtcctttt tggtggagga ggataaaaag cacgagcgcc acccaatctt tggcaatatc
     1081 gtggacgagg tggcgtacca tgaaaagtac ccaaccatat atcatctgag gaagaagctt
     1141 gtagacagta ctgataaggc tgacttgcgg ttgatctatc tcgcgctggc gcatatgatc
     1201 aaatttcggg gacacttcct catcgagggg gacctgaacc cagacaacag cgatgtcgac
     1261 aaactcttta tccaactggt tcagacttac aatcagcttt tcgaagagaa cccgatcaac
     1321 gcatccggag ttgacgccaa agcaatcctg agcgctaggc tgtccaaatc ccggcggctc
     1381 gaaaacctca tcgcacagct ccctggggag aagaagaacg gcctgtttgg taatcttatc
     1441 gccctgtcac tcgggctgac ccccaacttt aaatctaact tcgacctggc cgaagatgcc
     1501 aagcttcaac tgagcaaaga cacctacgat gatgatctcg acaatctgct ggcccagatc
     1561 ggcgaccagt acgcagacct ttttttggcg gcaaagaacc tgtcagacgc cattctgctg
     1621 agtgatattc tgcgagtgaa cacggagatc accaaagctc cgctgagcgc tagtatgatc
     1681 aagcgctatg atgagcacca ccaagacttg actttgctga aggcccttgt cagacagcaa
     1741 ctgcctgaga agtacaagga aattttcttc gatcagtcta aaaatggcta cgccggatac
     1801 attgacggcg gagcaagcca ggaggaattt tacaaattta ttaagcccat cttggaaaaa
     1861 atggacggca ccgaggagct gctggtaaag cttaacagag aagatctgtt gcgcaaacag
     1921 cgcactttcg acaatggaag catcccccac cagattcacc tgggcgaact gcacgctatc
     1981 ctcaggcggc aagaggattt ctaccccttt ttgaaagata acagggaaaa gattgagaaa
     2041 atcctcacat ttcggatacc ctactatgta ggccccctcg cccggggaaa ttccagattc
     2101 gcgtggatga ctcgcaaatc agaagagacc atcactccct ggaacttcga ggaagtcgtg
     2161 gataaggggg cctctgccca gtccttcatc gaaaggatga ctaactttga taaaaatctg
     2221 cctaacgaaa aggtgcttcc taaacactct ctgctgtacg agtacttcac agtttataac
     2281 gagctcacca aggtcaaata cgtcacagaa gggatgagaa agccagcatt cctgtctgga
     2341 gagcagaaga aagctatcgt ggacctcctc ttcaagacga accggaaagt taccgtgaaa
     2401 cagctcaaag aagactattt caaaaagatt gaatgtttcg actctgttga aatcagcgga
     2461 gtggaggatc gcttcaacgc atccctggga acgtatcacg atctcctgaa aatcattaaa
     2521 gacaaggact tcctggacaa tgaggagaac gaggacattc ttgaggacat tgtcctcacc
     2581 cttacgttgt ttgaagatag ggagatgatt gaagaacgct tgaaaactta cgctcatctc
     2641 ttcgacgaca aagtcatgaa acagctcaag aggcgccgat atacaggatg ggggcggctg
     2701 tcaagaaaac tgatcaatgg gatccgagac aagcagagtg gaaagacaat cctggatttt
     2761 cttaagtccg atggatttgc caaccggaac ttcatgcagt tgatccatga tgactctctc
     2821 acctttaagg aggacatcca gaaagcacaa gtttctggcc agggggacag tcttcacgag
     2881 cacatcgcta atcttgcagg tagcccagct atcaaaaagg gaatactgca gaccgttaag
     2941 gtcgtggatg aactcgtcaa agtaatggga aggcataagc ccgagaatat cgttatcgag
     3001 atggcccgag agaaccaaac tacccagaag ggacagaaga acagtaggga aaggatgaag
     3061 aggattgaag agggtataaa agaactgggg tcccaaatcc ttaaggaaca cccagttgaa
     3121 aacacccagc ttcagaatga gaagctctac ctgtactacc tgcagaacgg cagggacatg
     3181 tacgtggatc aggaactgga catcaatcgg ctctccgact acgacgtgga tcatatcgtg
     3241 ccccagtctt ttctcaaaga tgattctatt gataataaag tgttgacaag atccgataaa
     3301 aatagaggga agagtgataa cgtcccctca gaagaagttg tcaagaaaat gaaaaattat
     3361 tggcggcagc tgctgaacgc caaactgatc acacaacgga agttcgataa tctgactaag
     3421 gctgaacgag gtggcctgtc tgagttggat aaagccggct tcatcaaaag gcagcttgtt
     3481 gagacacgcc agatcaccaa gcacgtggcc caaattctcg attcacgcat gaacaccaag
     3541 tacgatgaaa atgacaaact gattcgagag gtgaaagtta ttactctgaa gtctaagctg
     3601 gtctcagatt tcagaaagga ctttcagttt tataaggtga gagagatcaa caattaccac
     3661 catgcgcatg atgcctacct gaatgcagtg gtaggcactg cacttatcaa aaaatatccc
     3721 aagcttgaat ctgaatttgt ttacggagac tataaagtgt acgatgttag gaaaatgatc
     3781 gcaaagtctg agcaggaaat aggcaaggcc accgctaagt acttctttta cagcaatatt
     3841 atgaattttt tcaagaccga gattacactg gccaatggag agattcggaa gcgaccactt
     3901 atcgaaacaa acggagaaac aggagaaatc gtgtgggaca agggtaggga tttcgcgaca
     3961 gtccggaagg tcctgtccat gccgcaggtg aacatcgtta aaaagaccga agtacagacc
     4021 ggaggcttct ccaaggaaag tatcctcccg aaaaggaaca gcgacaagct gatcgcacgc
     4081 aaaaaagatt gggaccccaa gaaatacggc ggattcgatt ctcctacagt cgcttacagt
     4141 gtactggttg tggccaaagt ggagaaaggg aagtctaaaa aactcaaaag cgtcaaggaa
     4201 ctgctgggca tcacaatcat ggagcgatca agcttcgaaa aaaaccccat cgactttctc
     4261 gaggcgaaag gatataaaga ggtcaaaaaa gacctcatca ttaagcttcc caagtactct
     4321 ctctttgagc ttgaaaacgg ccggaaacga atgctcgcta gtgcgggcga gctgcagaaa
     4381 ggtaacgagc tggcactgcc ctctaaatac gttaatttct tgtatctggc cagccactat
     4441 gaaaagctca aagggtctcc cgaagataat gagcagaagc agctgttcgt ggaacaacac
     4501 aaacactacc ttgatgagat catcgagcaa ataagcgaat tctccaaaag agtgatcctc
     4561 gccgacgcta acctcgataa ggtgctttct gcttacaata agcacaggga taagcccatc
     4621 agggagcagg cagaaaacat tatccacttg tttactctga ccaacttggg cgcgcctgca
     4681 gccttcaagt acttcgacac caccatagac agaaagcggt acacctctac aaaggaggtc
     4741 ctggacgcca cactgattca tcagtcaatt acggggctct atgaaacaag aatcgacctc
     4801 tctcagctcg gtggagacag cagggctgac cccaagaaga agaggaaggt gtgaaagggt
     4861 tcgatcccta ccggttagta atgagtttaa acgggggagg ctaactgaaa cacggaagga
     4921 gacaataccg gaaggaaccc gcgctatgac ggcaataaaa agacagaata aaacgcacgg
     4981 gtgttgggtc gtttgttcat aaacgcgggg ttcggtccca gggctggcac tctgtcgata
     5041 ccccaccgag accccattgg ggccaatacg cccgcgtttc ttccttttcc ccaccccacc
     5101 ccccaagttc gggtgaaggc ccagggctcg cagccaacgt cggggcggca ggccctgcca
     5161 tagcagatct gcgaaatcgg atccgcggcc gcactgattc tagctgcaga ggctccggtg
     5221 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
     5281 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
     5341 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
     5401 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggatcctcga gccaccatgg
     5461 ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg
     5521 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc
     5581 tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca
     5641 tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt
     5701 ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga
     5761 ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg
     5821 gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct
     5881 cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga
     5941 agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc
     6001 ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg
     6061 aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca
     6121 tggacgagct gtacaagtaa gttaacttgt ttattgcagc ttataatggt tacaaataaa
     6181 gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt
     6241 tgtccaaact catcaatgta tcttatcatg tctggaattg actcaaatga tgtcaattag
     6301 tctatcagaa gctcatctgg tctcccttcc gggggacaag acatccctgt ttaatattta
     6361 aacagcagtg ttcccaaact gggttcttat atcccttgct ctggtcaacc aggttgcagg
     6421 gtttcctgtc ctcacaggaa cgaagtccct aaagaaacag tggcagccag gtttagcccc
     6481 ggaattgact ggattccttt tttagggccc attggtatgg ctttttcccc gtatcccccc
     6541 aggtgtctgc aggctcaaag agcagcgaga agcgttcaga ggaaagcgat cccgtgccac
     6601 cttccccgtg cccgggctgt ccccgcacgc tgccggctcg gggatgcggg gggagcgccg
     6661 gaccggagcg gagccccggg cggctcgctg ctgcccccta gcgggggagg gacgtaatta
     6721 catccctggg ggctttgggg gggggctgtc cctgatatct ataacaagaa aatatatata
     6781 taataagtta tcacgtaagt agaacatgaa ataacaatat aattatcgta tgagttaaat
     6841 cttaaaagtc acgtaaaaga taatcatgcg tcattttgac tcacgcggtc gttatagttc
     6901 aaaatcagtg acacttaccg cattgacaag cacgcctcac gggagctcca agcggcgact
     6961 gagatgtcct aaatgcacag cgacggattc gcgctattta gaaagagaga gcaatatttc
     7021 aagaatgcat gcgtcaattt tacgcagact atctttctag ggttaatcta gctgcatcag
     7081 gatcatatcg tcgggtcttt tttccggctc agtcatcgcc caagctggcg ctatctgggc
     7141 atcggggagg aagaagcccg tgccttttcc cgcgaggttg aagcggcatg gaaagagttt
     7201 gccgaggatg actgctgctg cattgacgtt gagcgaaaac gcacgtttac catgatgatt
     7261 cgggaaggtg tggccatgca cgcctttaac ggtgaactgt tcgttcaggc cacctgggat
     7321 accagttcgt cgcggctttt ccggacacag ttccggatgg tcagcccgaa gcgcatcagc
     7381 aacccgaaca ataccggcga cagccggaac tgccgtgccg gtgtgcagat taatgacagc
     7441 ggtgcggcgc tgggatatta cgtcagcgag gacgggtatc ctggctggat gccgcagaaa
     7501 tggacatgga taccccgtga gttacccggc gggcgcgctt ggcgtaatca tggtcatagc
     7561 tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca
     7621 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     7681 cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     7741 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     7801 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     7861 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     7921 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     7981 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     8041 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     8101 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     8161 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     8221 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     8281 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     8341 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     8401 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     8461 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     8521 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     8581 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     8641 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     8701 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     8761 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     8821 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     8881 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     8941 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     9001 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     9061 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     9121 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     9181 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     9241 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     9301 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     9361 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     9421 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     9481 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     9541 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     9601 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     9661 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg taagcgttaa
     9721 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     9781 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     9841 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     9901 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     9961 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
    10021 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
    10081 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
    10141 tgcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt gggaagggcg
    10201 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
    10261 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga
    10321 gcgcgcctcg ttcattcacg tttttgaacc cgtggaggac gggcagactc gcggtgcaaa
    10381 tgtgttttac agcgtgatgg agcagatgaa gatgctcgac acgctgcaga acacgcagct
    10441 agattaaccc tagaaagata atcatattgt gacgtacgtt aaagataatc atgtgtaaaa
    10501 ttgacgcatg tgttttatcg gtctgtatat cgaggtttat ttattaattt gaatagatat
    10561 taagttttat tatatttaca cttacatact aataataaat tcaacaaaca atttatttat
    10621 gtttatttat ttattaaaaa aaacaaaaac tcaaaatttc ttctataaag taacaaaact
    10681 tttatgaggg acagcccccc cccaaagccc ccagggatgt aattacgtcc ctcccccgct
    10741 agggggcagc agcgagccgc ccggggctcc gctccggtcc ggcgctcccc ccgcatcccc
    10801 gagccggcag cgtgcgggga cagcccgggc acggggaagg tggcacggga tcgctttcct
    10861 ctgaacgctt ctcgctgctc tttgagcctg cagacacctg gggggatacg gggaaaaggc
    10921 ctccacggcc gacattgatt attgactagt tattaatagt aatcaattac ggggtcatta
    10981 gttcatagcc catatatgga gttccgcgtt acataactta cggtaaatgg cccgcctggc
    11041 tgaccgccca acgacccccg cccattgacg tcaataatga cgtatgttcc catagtaacg
    11101 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
    11161 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
    11221 tggcccgcct ggcattatgc ccagtacatg accttatggg actttcctac ttggcagtac
    11281 atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact
    11341 ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt
    11401 tgtgcagcga tgggggcggg gggggggggg gggsgsgcsc nggggggggg gggggggggg
    11461 gggggggggg gggggggggg gggggggggg gggggggggg gggngggggg gggggggggs
    11521 gggggggggg gggggggggg gggggggggg gggggggggg ggggg
//
