LOCUS       sequence-354043-       12295 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 12295)
  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 12295)
  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_Hygro
FEATURES             Location/Qualifiers
     source          1..12295
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          113..1129
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1053..1075
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1137..1156
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1225..1234
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1231..5334
                     /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             1231..5334
                     /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             1231..5334
                     /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             5347..5367
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5422..5441)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5466..5514
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        5746..5957
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      5967..5976
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5988..7007
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     CDS             5988..7007
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGYV
                     LRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPET
                     ELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQTV
                     MDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFGD
                     SQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGNF
                     DDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE"
     polyA_signal    7011..7132
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7048..7067)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7102..7121
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   7895..7929
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(8420..8436)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8420..8436)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8433..8455)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8444..8460
                     /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(8468..8474,8475..8492,8493..8498))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8513..8534
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8651..8668)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8822..9410)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8902..8921)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9581..10372,10373..10441))
                     /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     10204..10223
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10442..10546)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(10572..11027)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10709..10730)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     10921..10940
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11154..11176
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     11168..11185
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     11169..11185
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   11315..11377
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        11798..12177
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt ttccttttat
       61 ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc
      121 tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct
      181 ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc ctccgggctg
      241 taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg aaagccttga
      301 ggggctccgg gagggccctt tgtgcggggg gagcggctcg gggggtgcgt gcgtgtgtgt
      361 gtgcgtgggg agcgccgcgt gcggctccgc gctgcccggc ggctgtgagc gctgcgggcg
      421 cggcgcgggg ctttttgcgc tccgcagtgt gcgcgagggg agcgcggccg ggggcggtgc
      481 cccgcggtgc ggggggggct gcgaggggaa caaaggctgc gtgcggggtg tgtgcgtggg
      541 ggggtgagca gggggtgtgg gcgcgtcggt cgggctgcaa ccccccctgc acccccctcc
      601 ccgagttgct gagcacggcc cggcttcggg tgcggggctc cgtacggggc gtggcgcggg
      661 gctcgccgtg ccgggcgggg ggtggcggca ggtgggggtg ccgggcgggg cggggccgcc
      721 tcgggccggg gagggctcgg gggaggggcg cggcggcccc cggagcgccg gcggctgtcg
      781 aggcgcggcg agccgcagcc attgcctttt atggtaatcg tgcgagaggg cgcagggact
      841 tcctttgtcc caaatctgtg cggagccgaa atctgggagg cgccgccgca ccccctctag
      901 cgggcgcggg gcgaagcggt gcggcgccgg caggaaggaa atgggcgggg agggccttcg
      961 tgcgtcgccg cgccgccgtc cccttctccc tctccagcct cggggctgtc cgcgggggga
     1021 cggctgcctt cgggggggac ggggcagggc ggggttcggc ttctggcgtg tgaccggcgg
     1081 ctctagagcc tctgctaacc atgttcatgc cttcttcttt ttcctacagc tcctgggcaa
     1141 cgtgctggtt attgtgctgt ctcatcattt tggcaaagaa tctagatacc ctcgtaaagg
     1201 tctagagcta gcgaattcga atttgccacc atggacaaga agtactccat tgggctcgat
     1261 atcggcacaa acagcgtcgg ctgggccgtc attacggacg agtacaaggt gccgagcaaa
     1321 aaattcaaag ttctgggcaa taccgatcgc cacagcataa agaagaacct cattggcgcc
     1381 ctcctgttcg actccgggga gacggccgaa gccacgcggc tcaaaagaac agcacggcgc
     1441 agatataccc gcagaaagaa tcggatctgc tacctgcagg agatctttag taatgagatg
     1501 gctaaggtgg atgactcttt cttccatagg ctggaggagt cctttttggt ggaggaggat
     1561 aaaaagcacg agcgccaccc aatctttggc aatatcgtgg acgaggtggc gtaccatgaa
     1621 aagtacccaa ccatatatca tctgaggaag aagcttgtag acagtactga taaggctgac
     1681 ttgcggttga tctatctcgc gctggcgcat atgatcaaat ttcggggaca cttcctcatc
     1741 gagggggacc tgaacccaga caacagcgat gtcgacaaac tctttatcca actggttcag
     1801 acttacaatc agcttttcga agagaacccg atcaacgcat ccggagttga cgccaaagca
     1861 atcctgagcg ctaggctgtc caaatcccgg cggctcgaaa acctcatcgc acagctccct
     1921 ggggagaaga agaacggcct gtttggtaat cttatcgccc tgtcactcgg gctgaccccc
     1981 aactttaaat ctaacttcga cctggccgaa gatgccaagc ttcaactgag caaagacacc
     2041 tacgatgatg atctcgacaa tctgctggcc cagatcggcg accagtacgc agaccttttt
     2101 ttggcggcaa agaacctgtc agacgccatt ctgctgagtg atattctgcg agtgaacacg
     2161 gagatcacca aagctccgct gagcgctagt atgatcaagc gctatgatga gcaccaccaa
     2221 gacttgactt tgctgaaggc ccttgtcaga cagcaactgc ctgagaagta caaggaaatt
     2281 ttcttcgatc agtctaaaaa tggctacgcc ggatacattg acggcggagc aagccaggag
     2341 gaattttaca aatttattaa gcccatcttg gaaaaaatgg acggcaccga ggagctgctg
     2401 gtaaagctta acagagaaga tctgttgcgc aaacagcgca ctttcgacaa tggaagcatc
     2461 ccccaccaga ttcacctggg cgaactgcac gctatcctca ggcggcaaga ggatttctac
     2521 ccctttttga aagataacag ggaaaagatt gagaaaatcc tcacatttcg gataccctac
     2581 tatgtaggcc ccctcgcccg gggaaattcc agattcgcgt ggatgactcg caaatcagaa
     2641 gagaccatca ctccctggaa cttcgaggaa gtcgtggata agggggcctc tgcccagtcc
     2701 ttcatcgaaa ggatgactaa ctttgataaa aatctgccta acgaaaaggt gcttcctaaa
     2761 cactctctgc tgtacgagta cttcacagtt tataacgagc tcaccaaggt caaatacgtc
     2821 acagaaggga tgagaaagcc agcattcctg tctggagagc agaagaaagc tatcgtggac
     2881 ctcctcttca agacgaaccg gaaagttacc gtgaaacagc tcaaagaaga ctatttcaaa
     2941 aagattgaat gtttcgactc tgttgaaatc agcggagtgg aggatcgctt caacgcatcc
     3001 ctgggaacgt atcacgatct cctgaaaatc attaaagaca aggacttcct ggacaatgag
     3061 gagaacgagg acattcttga ggacattgtc ctcaccctta cgttgtttga agatagggag
     3121 atgattgaag aacgcttgaa aacttacgct catctcttcg acgacaaagt catgaaacag
     3181 ctcaagaggc gccgatatac aggatggggg cggctgtcaa gaaaactgat caatgggatc
     3241 cgagacaagc agagtggaaa gacaatcctg gattttctta agtccgatgg atttgccaac
     3301 cggaacttca tgcagttgat ccatgatgac tctctcacct ttaaggagga catccagaaa
     3361 gcacaagttt ctggccaggg ggacagtctt cacgagcaca tcgctaatct tgcaggtagc
     3421 ccagctatca aaaagggaat actgcagacc gttaaggtcg tggatgaact cgtcaaagta
     3481 atgggaaggc ataagcccga gaatatcgtt atcgagatgg cccgagagaa ccaaactacc
     3541 cagaagggac agaagaacag tagggaaagg atgaagagga ttgaagaggg tataaaagaa
     3601 ctggggtccc aaatccttaa ggaacaccca gttgaaaaca cccagcttca gaatgagaag
     3661 ctctacctgt actacctgca gaacggcagg gacatgtacg tggatcagga actggacatc
     3721 aatcggctct ccgactacga cgtggatcat atcgtgcccc agtcttttct caaagatgat
     3781 tctattgata ataaagtgtt gacaagatcc gataaaaata gagggaagag tgataacgtc
     3841 ccctcagaag aagttgtcaa gaaaatgaaa aattattggc ggcagctgct gaacgccaaa
     3901 ctgatcacac aacggaagtt cgataatctg actaaggctg aacgaggtgg cctgtctgag
     3961 ttggataaag ccggcttcat caaaaggcag cttgttgaga cacgccagat caccaagcac
     4021 gtggcccaaa ttctcgattc acgcatgaac accaagtacg atgaaaatga caaactgatt
     4081 cgagaggtga aagttattac tctgaagtct aagctggtct cagatttcag aaaggacttt
     4141 cagttttata aggtgagaga gatcaacaat taccaccatg cgcatgatgc ctacctgaat
     4201 gcagtggtag gcactgcact tatcaaaaaa tatcccaagc ttgaatctga atttgtttac
     4261 ggagactata aagtgtacga tgttaggaaa atgatcgcaa agtctgagca ggaaataggc
     4321 aaggccaccg ctaagtactt cttttacagc aatattatga attttttcaa gaccgagatt
     4381 acactggcca atggagagat tcggaagcga ccacttatcg aaacaaacgg agaaacagga
     4441 gaaatcgtgt gggacaaggg tagggatttc gcgacagtcc ggaaggtcct gtccatgccg
     4501 caggtgaaca tcgttaaaaa gaccgaagta cagaccggag gcttctccaa ggaaagtatc
     4561 ctcccgaaaa ggaacagcga caagctgatc gcacgcaaaa aagattggga ccccaagaaa
     4621 tacggcggat tcgattctcc tacagtcgct tacagtgtac tggttgtggc caaagtggag
     4681 aaagggaagt ctaaaaaact caaaagcgtc aaggaactgc tgggcatcac aatcatggag
     4741 cgatcaagct tcgaaaaaaa ccccatcgac tttctcgagg cgaaaggata taaagaggtc
     4801 aaaaaagacc tcatcattaa gcttcccaag tactctctct ttgagcttga aaacggccgg
     4861 aaacgaatgc tcgctagtgc gggcgagctg cagaaaggta acgagctggc actgccctct
     4921 aaatacgtta atttcttgta tctggccagc cactatgaaa agctcaaagg gtctcccgaa
     4981 gataatgagc agaagcagct gttcgtggaa caacacaaac actaccttga tgagatcatc
     5041 gagcaaataa gcgaattctc caaaagagtg atcctcgccg acgctaacct cgataaggtg
     5101 ctttctgctt acaataagca cagggataag cccatcaggg agcaggcaga aaacattatc
     5161 cacttgttta ctctgaccaa cttgggcgcg cctgcagcct tcaagtactt cgacaccacc
     5221 atagacagaa agcggtacac ctctacaaag gaggtcctgg acgccacact gattcatcag
     5281 tcaattacgg ggctctatga aacaagaatc gacctctctc agctcggtgg agacagcagg
     5341 gctgacccca agaagaagag gaaggtgtga aagggttcga tccctaccgg ttagtaatga
     5401 gtttaaacgg gggaggctaa ctgaaacacg gaaggagaca ataccggaag gaacccgcgc
     5461 tatgacggca ataaaaagac agaataaaac gcacgggtgt tgggtcgttt gttcataaac
     5521 gcggggttcg gtcccagggc tggcactctg tcgatacccc accgagaccc cattggggcc
     5581 aatacgcccg cgtttcttcc ttttccccac cccacccccc aagttcgggt gaaggcccag
     5641 ggctcgcagc caacgtcggg gcggcaggcc ctgccatagc agatctgcga aatcggatcc
     5701 gcggccgcac tgattctagc tgcagaggct ccggtgcccg tcagtgggca gagcgcacat
     5761 cgcccacagt ccccgagaag ttggggggag gggtcggcaa ttgaaccggt gcctagagaa
     5821 ggtggcgcgg ggtaaactgg gaaagtgatg tcgtgtactg gctccgcctt tttcccgagg
     5881 gtgggggaga accgtatata agtgcagtag tcgccgtgaa cgttcttttt cgcaacgggt
     5941 ttgccgccag aacacaggat cctcgagcca ccatggatag atccggaaag cctgaactca
     6001 ccgcgacgtc tgtcgagaag tttctgatcg aaaagttcga cagcgtttcc gacctgatgc
     6061 agctctcgga gggcgaagaa tctcgtgctt tcagcttcga tgtaggaggg cgtggatatg
     6121 tcctgcgggt aaatagctgc gccgatggtt tctacaaaga tcgttatgtt tatcggcact
     6181 ttgcatcggc cgcgctcccg attccggaag tgcttgacat tggggaattc agcgagagcc
     6241 tgacctattg catctcccgc cgtgcacagg gtgtcacgtt gcaagacctg cctgaaaccg
     6301 aactgcccgc tgttctgcag ccggtcgcgg aggccatgga tgcgatcgct gcggccgatc
     6361 ttagccagac gagcgggttc ggcccattcg gaccgcaagg aatcggtcaa tacactacat
     6421 ggcgtgattt catatgcgcg attgctgatc cccatgtgta tcactggcaa actgtgatgg
     6481 acgacaccgt cagtgcgtcc gtcgcgcagg ctctcgatga gctgatgctt tgggccgagg
     6541 actgccccga agtccggcac ctcgtgcacg cggatttcgg ctccaacaat gtcctgacgg
     6601 acaatggccg cataacagcg gtcattgact ggagcgaggc gatgttcggg gattcccaat
     6661 acgaggtcgc caacatcttc ttctggaggc cgtggttggc ttgtatggag cagcagacgc
     6721 gctacttcga gcggaggcat ccggagcttg caggatcgcc gcggctccgg gcgtatatgc
     6781 tccgcattgg tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag
     6841 cttgggcgca gggtcgatgc gacgcaatcg tccgatccgg agccgggact gtcgggcgta
     6901 cacaaatcgc ccgcagaagc gcggccgtct ggaccgatgg ctgtgtagaa gtactcgccg
     6961 atagtggaaa ccgacgcccc agcactcgtc cgagggcaaa ggaataggtt aacttgttta
     7021 ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca aataaagcat
     7081 ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct tatcatgtct
     7141 ggaattgact caaatgatgt caattagtct atcagaagct catctggtct cccttccggg
     7201 ggacaagaca tccctgttta atatttaaac agcagtgttc ccaaactggg ttcttatatc
     7261 ccttgctctg gtcaaccagg ttgcagggtt tcctgtcctc acaggaacga agtccctaaa
     7321 gaaacagtgg cagccaggtt tagccccgga attgactgga ttcctttttt agggcccatt
     7381 ggtatggctt tttccccgta tccccccagg tgtctgcagg ctcaaagagc agcgagaagc
     7441 gttcagagga aagcgatccc gtgccacctt ccccgtgccc gggctgtccc cgcacgctgc
     7501 cggctcgggg atgcgggggg agcgccggac cggagcggag ccccgggcgg ctcgctgctg
     7561 ccccctagcg ggggagggac gtaattacat ccctgggggc tttggggggg ggctgtccct
     7621 gatatctata acaagaaaat atatatataa taagttatca cgtaagtaga acatgaaata
     7681 acaatataat tatcgtatga gttaaatctt aaaagtcacg taaaagataa tcatgcgtca
     7741 ttttgactca cgcggtcgtt atagttcaaa atcagtgaca cttaccgcat tgacaagcac
     7801 gcctcacggg agctccaagc ggcgactgag atgtcctaaa tgcacagcga cggattcgcg
     7861 ctatttagaa agagagagca atatttcaag aatgcatgcg tcaattttac gcagactatc
     7921 tttctagggt taatctagct gcatcaggat catatcgtcg ggtctttttt ccggctcagt
     7981 catcgcccaa gctggcgcta tctgggcatc ggggaggaag aagcccgtgc cttttcccgc
     8041 gaggttgaag cggcatggaa agagtttgcc gaggatgact gctgctgcat tgacgttgag
     8101 cgaaaacgca cgtttaccat gatgattcgg gaaggtgtgg ccatgcacgc ctttaacggt
     8161 gaactgttcg ttcaggccac ctgggatacc agttcgtcgc ggcttttccg gacacagttc
     8221 cggatggtca gcccgaagcg catcagcaac ccgaacaata ccggcgacag ccggaactgc
     8281 cgtgccggtg tgcagattaa tgacagcggt gcggcgctgg gatattacgt cagcgaggac
     8341 gggtatcctg gctggatgcc gcagaaatgg acatggatac cccgtgagtt acccggcggg
     8401 cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     8461 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     8521 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     8581 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     8641 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     8701 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     8761 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     8821 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     8881 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     8941 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     9001 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     9061 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     9121 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     9181 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     9241 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     9301 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     9361 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     9421 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     9481 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     9541 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     9601 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     9661 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     9721 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     9781 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     9841 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     9901 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     9961 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
    10021 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
    10081 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
    10141 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
    10201 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
    10261 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
    10321 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
    10381 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
    10441 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
    10501 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
    10561 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
    10621 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
    10681 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
    10741 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
    10801 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
    10861 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
    10921 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
    10981 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
    11041 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
    11101 ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt tttcccagtc
    11161 acgacgttgt aaaacgacgg ccagtgagcg cgcctcgttc attcacgttt ttgaacccgt
    11221 ggaggacggg cagactcgcg gtgcaaatgt gttttacagc gtgatggagc agatgaagat
    11281 gctcgacacg ctgcagaaca cgcagctaga ttaaccctag aaagataatc atattgtgac
    11341 gtacgttaaa gataatcatg tgtaaaattg acgcatgtgt tttatcggtc tgtatatcga
    11401 ggtttattta ttaatttgaa tagatattaa gttttattat atttacactt acatactaat
    11461 aataaattca acaaacaatt tatttatgtt tatttattta ttaaaaaaaa caaaaactca
    11521 aaatttcttc tataaagtaa caaaactttt atgagggaca gccccccccc aaagccccca
    11581 gggatgtaat tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct
    11641 ccggtccggc gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg
    11701 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
    11761 acacctgggg ggatacgggg aaaaggcctc cacggccgac attgattatt gactagttat
    11821 taatagtaat caattacggg gtcattagtt catagcccat atatggagtt ccgcgttaca
    11881 taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca
    11941 ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg
    12001 gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg
    12061 ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc
    12121 ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc
    12181 gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt
    12241 ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg ggggg
//
