LOCUS       sequence-273623-        9207 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of mTN3-GGS6-dCas9 (iCas9) in human..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9207)
  AUTHORS   Standage-Beier K, Brookhouser N, Balachandran P, Zhang Q, Brafman 
            DA, Wang X
  TITLE     RNA-Guided Recombinase-Cas9 Fusion Targets Genomic DNA Deletion and 
            Integration.
  JOURNAL   CRISPR J. 2019 Aug;2:209-222. doi: 10.1089/crispr.2019.0013.
  PUBMED    31436506
REFERENCE   2  (bases 1 to 9207)
  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: "CRISPR J.";
            date: "2019-08"; volume: "2:209-222. doi"; pages: " 
            10.1089/crispr.2019.0013"
COMMENT     Sequence Label: pEF:iCas9
FEATURES             Location/Qualifiers
     source          1..9207
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          221..1163
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1120..1140
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             1240..1260
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1771..1794,1795..1797,1798..4281,4282..4287,4288..4353,
                     4354..4356,4357..5871)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=Cas9m4
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the mutations D10A, D839A, 
                     H840A, and N863A (Mali et al., 2013)"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             join(1771..1794,1795..1797,1798..4284,4285..4287,
                     4288..5871)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1771..5871
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5884..5904
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5936..5954)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(6183..6202)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    6248..6303
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(6302..6321)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(6664..6680)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6664..6680)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6677..6699)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6688..6704
                     /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(6712..6718,6719..6736,6737..6742))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6757..6778
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        6837..7032
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      6883..7018
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    7038..7172
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7075..7094)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7129..7148
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(7240..7257)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7411..7999)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7491..7510)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8170..8961,8962..9030))
                     /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     8793..8812
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9031..9135)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        join(9197..9207,1..1172)
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
ORIGIN
        1 ccgtcagtgg gcagagcgca catcgcccac agtccccgag aagttggggg gaggggtcgg
       61 caattgaacc ggtgcctaga gaaggtggcg cggggtaaac tgggaaagtg atgtcgtgta
      121 ctggctccgc ctttttcccg agggtggggg agaaccgtat ataagtgcag tagtcgccgt
      181 gaacgttctt tttcgcaacg ggtttgccgc cagaacacag gtaagtgccg tgtgtggttc
      241 ccgcgggcct ggcctcttta cgggttatgg cccttgcgtg ccttgaatta cttccacgcc
      301 cctggctgca gtacgtgatt cttgatcccg agcttcgggt tggaagtggg tgggagagtt
      361 cgaggccttg cgcttaagga gccccttcgc ctcgtgcttg agttgaggcc tggcctgggc
      421 gctggggccg ccgcgtgcga atctggtggc accttcgcgc ctgtctcgct gctttcgata
      481 agtctctagc catttaaaat ttttgatgac ctgctgcgac gctttttttc tggcaagata
      541 gtcttgtaaa tgcgggccaa gatctgcaca ctggtatttc ggtttttggg gccgcgggcg
      601 gcgacggggc ccgtgcgtcc cagcgcacat gttcggcgag gcggggcctg cgagcgcggc
      661 caccgagaat cggacggggg tagtctcaag ctggccggcc tgctctggtg cctggcctcg
      721 cgccgccgtg tatcgccccg ccctgggcgg caaggctggc ccggtcggca ccagttgcgt
      781 gagcggaaag atggccgctt cccggccctg ctgcagggag ctcaaaatgg aggacgcggc
      841 gctcgggaga gcgggcgggt gagtcaccca cacaaaggaa aagggccttt ccgtcctcag
      901 ccgtcgcttc atgtgactcc acggagtacc gggcgccgtc caggcacctc gattagttct
      961 cgagcttttg gagtacgtcg tctttaggtt ggggggaggg gttttatgcg atggagtttc
     1021 cccacactga gtgggtggag actgaagtta ggccagcttg gcacttgatg taattctcct
     1081 tggaatttgc cctttttgag tttggatctt ggttcattct caagcctcag acagtggttc
     1141 aaagtttttt tcttccattt caggtgtcgt gaggaattcg agctcggtac ccggggatcc
     1201 tctagaacta gaactagtgg atcccccggg aaaaaaatgc ccaaaaagaa gaggaaagta
     1261 ggtggcagtg caaggatatt tggatacgca cgagttagta catctcaaca gtctctggac
     1321 atacagataa gggcattgaa ggatgctgga gtgaaagcta atcggatctt taccgataag
     1381 gctagtggca gctctacaga cagagagggc ttggatctgc tgagaatgaa ggttgaggaa
     1441 ggggatgtga tactcgtcaa gaaacttgac agactctcac gagacactgc cgatatgatc
     1501 caactgatta aggaatttga cgcccaaggt gtagccgtaa gatttattga tgacggtatc
     1561 agcaccgacg gctatatggg ccagatggtc gtcactatac tgagcgctgt cgcacaagcg
     1621 gagagaagac gaatcctgca acggacaaat gaaggtcgcc aagaagcaaa gctcaaagga
     1681 attaagttcg ggcgcaggag gactgtagac agaggtggtt caggaggaag tggaggttca
     1741 gggggaagtg gtggctccgg tgggtctgca gacaagaagt actccattgg gctcgccatc
     1801 ggcacaaaca gcgtcggttg ggccgtcatt acggacgagt acaaggtgcc gagcaaaaaa
     1861 ttcaaagttc tgggcaatac cgatcgccac agcataaaga agaacctcat tggcgccctc
     1921 ctgttcgact ccggggagac ggccgaagcc acgcggctca aaagaacagc acggcgcaga
     1981 tatacccgca gaaagaatcg gatctgctac ctgcaggaga tctttagtaa tgagatggct
     2041 aaggtggatg actctttctt ccataggctg gaggagtcct ttttggtgga ggaggataaa
     2101 aagcacgagc gccacccaat ctttggcaat atcgtggacg aggtggcgta ccatgaaaag
     2161 tacccaacca tatatcatct gaggaagaag cttgtagaca gtactgataa ggctgacttg
     2221 cggttgatct atctcgcgct ggcgcatatg atcaaatttc ggggacactt cctcatcgag
     2281 ggggacctga acccagacaa cagcgatgtc gacaaactct ttatccaact ggttcagact
     2341 tacaatcagc ttttcgaaga gaacccgatc aacgcatccg gagttgacgc caaagcaatc
     2401 ctgagcgcta ggctgtccaa atcccggcgg ctcgaaaacc tcatcgcaca gctccctggg
     2461 gagaagaaga acggcctgtt tggtaatctt atcgccctgt cactcgggct gacccccaac
     2521 tttaaatcta acttcgacct ggccgaagat gccaagcttc aactgagcaa agacacctac
     2581 gatgatgatc tcgacaatct gctggcccag atcggcgacc agtacgcaga cctttttttg
     2641 gcggcaaaga acctgtcaga cgccattctg ctgagtgata ttctgcgagt gaacacggag
     2701 atcaccaaag ctccgctgag cgctagtatg atcaagcgct atgatgagca ccaccaagac
     2761 ttgactttgc tgaaggccct tgtcagacag caactgcctg agaagtacaa ggaaattttc
     2821 ttcgatcagt ctaaaaatgg ctacgccgga tacattgacg gcggagcaag ccaggaggaa
     2881 ttttacaaat ttattaagcc catcttggaa aaaatggacg gcaccgagga gctgctggta
     2941 aagcttaaca gagaagatct gttgcgcaaa cagcgcactt tcgacaatgg aagcatcccc
     3001 caccagattc acctgggcga actgcacgct atcctcaggc ggcaagagga tttctacccc
     3061 tttttgaaag ataacaggga aaagattgag aaaatcctca catttcggat accctactat
     3121 gtaggccccc tcgcccgggg aaattccaga ttcgcgtgga tgactcgcaa atcagaagag
     3181 accatcactc cctggaactt cgaggaagtc gtggataagg gggcctctgc ccagtccttc
     3241 atcgaaagga tgactaactt tgataaaaat ctgcctaacg aaaaggtgct tcctaaacac
     3301 tctctgctgt acgagtactt cacagtttat aacgagctca ccaaggtcaa atacgtcaca
     3361 gaagggatga gaaagccagc attcctgtct ggagagcaga agaaagctat cgtggacctc
     3421 ctcttcaaga cgaaccggaa agttaccgtg aaacagctca aagaagacta tttcaaaaag
     3481 attgaatgtt tcgactctgt tgaaatcagc ggagtggagg atcgcttcaa cgcatccctg
     3541 ggaacgtatc acgatctcct gaaaatcatt aaagacaagg acttcctgga caatgaggag
     3601 aacgaggaca ttcttgagga cattgtcctc acccttacgt tgtttgaaga tagggagatg
     3661 attgaagaac gcttgaaaac ttacgctcat ctcttcgacg acaaagtcat gaaacagctc
     3721 aagaggcgcc gatatacagg atgggggcgg ctgtcaagaa aactgatcaa tgggatccga
     3781 gacaagcaga gtggaaagac aatcctggat tttcttaagt ccgatggatt tgccaaccgg
     3841 aacttcatgc agttgatcca tgatgactct ctcaccttta aggaggacat ccagaaagca
     3901 caagtttctg gccaggggga cagtcttcac gagcacatcg ctaatcttgc aggtagccca
     3961 gctatcaaaa agggaatact gcagaccgtt aaggtcgtgg atgaactcgt caaagtaatg
     4021 ggaaggcata agcccgagaa tatcgttatc gagatggccc gagagaacca aactacccag
     4081 aagggacaga agaacagtag ggaaaggatg aagaggattg aagagggtat aaaagaactg
     4141 gggtcccaaa tccttaagga acacccagtt gaaaacaccc agcttcagaa tgagaagctc
     4201 tacctgtact acctgcagaa cggcagggac atgtacgtgg atcaggaact ggacatcaat
     4261 cggctctccg actacgacgt ggatgccatc gtgccccagt cttttctcaa agatgattct
     4321 attgataata aagtgttgac aagatccgat aaaaatagag ggaagagtga taacgtcccc
     4381 tcagaagaag ttgtcaagaa aatgaaaaat tattggcggc agctgctgaa cgccaaactg
     4441 atcacacaac ggaagttcga taatctgact aaggctgaac gaggtggcct gtctgagttg
     4501 gataaagccg gcttcatcaa aaggcagctt gttgagacac gccagatcac caagcacgtg
     4561 gcccaaattc tcgattcacg catgaacacc aagtacgatg aaaatgacaa actgattcga
     4621 gaggtgaaag ttattactct gaagtctaag ctggtctcag atttcagaaa ggactttcag
     4681 ttttataagg tgagagagat caacaattac caccatgcgc atgatgccta cctgaatgca
     4741 gtggtaggca ctgcacttat caaaaaatat cccaagcttg aatctgaatt tgtttacgga
     4801 gactataaag tgtacgatgt taggaaaatg atcgcaaagt ctgagcagga aataggcaag
     4861 gccaccgcta agtacttctt ttacagcaat attatgaatt ttttcaagac cgagattaca
     4921 ctggccaatg gagagattcg gaagcgacca cttatcgaaa caaacggaga aacaggagaa
     4981 atcgtgtggg acaagggtag ggatttcgcg acagtccgga aggtcctgtc catgccgcag
     5041 gtgaacatcg ttaaaaagac cgaagtacag accggaggct tctccaagga aagtatcctc
     5101 ccgaaaagga acagcgacaa gctgatcgca cgcaaaaaag attgggaccc caagaaatac
     5161 ggcggattcg attctcctac agtcgcttac agtgtactgg ttgtggccaa agtggagaaa
     5221 gggaagtcta aaaaactcaa aagcgtcaag gaactgctgg gcatcacaat catggagcga
     5281 tcaagcttcg aaaaaaaccc catcgacttt ctcgaggcga aaggatataa agaggtcaaa
     5341 aaagacctca tcattaagct tcccaagtac tctctctttg agcttgaaaa cggccggaaa
     5401 cgaatgctcg ctagtgcggg cgagctgcag aaaggtaacg agctggcact gccctctaaa
     5461 tacgttaatt tcttgtatct ggccagccac tatgaaaagc tcaaagggtc tcccgaagat
     5521 aatgagcaga agcagctgtt cgtggaacaa cacaaacact accttgatga gatcatcgag
     5581 caaataagcg aattctccaa aagagtgatc ctcgccgacg ctaacctcga taaggtgctt
     5641 tctgcttaca ataagcacag ggataagccc atcagggagc aggcagaaaa cattatccac
     5701 ttgtttactc tgaccaactt gggcgcgcct gcagccttca agtacttcga caccaccata
     5761 gacagaaagc ggtacacctc tacaaaggag gtcctggacg ccacactgat tcatcagtca
     5821 attacggggc tctatgaaac aagaatcgac ctctctcagc tcggtggaga cagcagggct
     5881 gaccccaaga agaagaggaa ggtgtgatct cttctcgagt catgtaatta gttatgtcac
     5941 gcttacattc acgccctccc cccacatccg ctctaaccga aaaggaagga gttagacaac
     6001 ctgaagtcta ggtccctatt tattttttta tagttatgtt agtattaaga acgttattta
     6061 tatttcaaat ttttcttttt tttctgtaca gacgcgtgcg gccgcactcc tcaggtgcag
     6121 gctgcctatc agaaggtggt ggctggtgtg gccaatgccc tggctcacaa ataccactga
     6181 gatctttttc cctctgccaa aaattatggg gacatcatga agccccttga gcatctgact
     6241 tctggctaat aaaggaaatt tattttcatt gcaatagtgt gttggaattt tttgtgtctc
     6301 tcactcggaa ggacatatgg gagggcaaat catttaaaac atcagaatga gtatttggtt
     6361 tagagtttgg caacatatgc ccatatgctg gctgccatga acaaaggttg gctataaaga
     6421 ggtcatcagt atatgaaaca gccccctgct gtccattcct tattccatag aaaagccttg
     6481 acttgaggtt agattttttt tatattttgt tttgtgttat ttttttcttt aacatcccta
     6541 aaattttcct tacatgtttt actagccaga tttttcctcc tctcctgact actcccagtc
     6601 atagctgtcc ctcttctctt atggagatcc ctcgacctgc agcccaagct tggcgtaatc
     6661 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     6721 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     6781 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc ggatccgcat
     6841 ctcaattagt cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg
     6901 cccagttccg cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc
     6961 gaggccgcct cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta
     7021 ggcttttgca aaaagctaac ttgtttattg cagcttataa tggttacaaa taaagcaata
     7081 gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca
     7141 aactcatcaa tgtatcttat catgtctgga tccgctgcat taatgaatcg gccaacgcgc
     7201 ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc tcgctcactg actcgctgcg
     7261 ctcggtcgtt cggctgcggc gagcggtatc agctcactca aaggcggtaa tacggttatc
     7321 cacagaatca ggggataacg caggaaagaa catgtgagca aaaggccagc aaaaggccag
     7381 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     7441 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     7501 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     7561 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     7621 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     7681 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     7741 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     7801 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     7861 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     7921 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     7981 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     8041 gaacgaaaac tcacgttaag ggattttggt catgagatta tcaaaaagga tcttcaccta
     8101 gatcctttta aattaaaaat gaagttttaa atcaatctaa agtatatatg agtaaacttg
     8161 gtctgacagt taccaatgct taatcagtga ggcacctatc tcagcgatct gtctatttcg
     8221 ttcatccata gttgcctgac tccccgtcgt gtagataact acgatacggg agggcttacc
     8281 atctggcccc agtgctgcaa tgataccgcg agacccacgc tcaccggctc cagatttatc
     8341 agcaataaac cagccagccg gaagggccga gcgcagaagt ggtcctgcaa ctttatccgc
     8401 ctccatccag tctattaatt gttgccggga agctagagta agtagttcgc cagttaatag
     8461 tttgcgcaac gttgttgcca ttgctacagg catcgtggtg tcacgctcgt cgtttggtat
     8521 ggcttcattc agctccggtt cccaacgatc aaggcgagtt acatgatccc ccatgttgtg
     8581 caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc agaagtaagt tggccgcagt
     8641 gttatcactc atggttatgg cagcactgca taattctctt actgtcatgc catccgtaag
     8701 atgcttttct gtgactggtg agtactcaac caagtcattc tgagaatagt gtatgcggcg
     8761 accgagttgc tcttgcccgg cgtcaatacg ggataatacc gcgccacata gcagaacttt
     8821 aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa ctctcaagga tcttaccgct
     8881 gttgagatcc agttcgatgt aacccactcg tgcacccaac tgatcttcag catcttttac
     8941 tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa aatgccgcaa aaaagggaat
     9001 aagggcgaca cggaaatgtt gaatactcat actcttcctt tttcaatatt attgaagcat
     9061 ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga aaaataaaca
     9121 aataggggtt ccgcgcacat ttccccgaaa agtgccacct gggtcgacat tgattattga
     9181 ctagatcatc gcgtgaggct ccggtgc
//
