LOCUS       sequence-249095-        9826 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Construct for generating stable Cas9-expressing Toxoplasma gondii.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9826)
  AUTHORS   Markus BM, Bell GW, Lorenzi HA, Lourido S
  TITLE     Optimizing Systems for Cas9 Expression in Toxoplasma gondii.
  JOURNAL   mSphere. 2019 Jun 26;4(3). pii: 4/3/e00386-19. doi: 
            10.1128/mSphere.00386-19.
  PUBMED    31243081
REFERENCE   2  (bases 1 to 9826)
  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: "mSphere."; 
            date: "2019-06-26"; volume: "4(3). pii: 4/3/e00386-19. doi"; pages: 
            " 10.1128/mSphere.00386-19"
COMMENT     Sequence Label: pBM019
FEATURES             Location/Qualifiers
     source          1..9826
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        297..372
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     CDS             908..973
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             980..1000
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1025..5125
                     /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             1025..5125
                     /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             5126..5173
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5189..5245
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             5255..5914
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MHEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAF
                     LKTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFS
                     SLWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVAN
                     MDNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(5324..5343)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             5915..5941
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     promoter        complement(6719..6737)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(6758..6774)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6758..6774)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6771..6793)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6782..6798
                     /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(6806..6812,6813..6830,6831..6836))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6851..6872
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6989..7006)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7160..7748)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7240..7259)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7919..8710,8711..8779))
                     /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     8542..8561
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8780..8884)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(8910..9365)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9047..9068)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     9259..9278
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9492..9514
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     9506..9523
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     9507..9523
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9530..9549
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        9530..9548
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     9573..9589
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     9574..9590
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
ORIGIN
        1 ctgtgaattg ggcgccaata ttgcatccta ggcctgacgc gcctcctgca gaacgcgaga
       61 cactgggata tgtagagcca agggggaaac cttcgaactc tcgaatgtct tctctgacaa
      121 gaatcatatt tccatcagtt ctgtcagatt ttcaaatggc gacctgcaga ggcctgcttc
      181 ctccctgtgc gctcttcgaa ggggctttct gtcgcgcagg gtcacctcgt ccccgaaggg
      241 ggtgtttgcc ttctggtaaa tggggatgtc aagttgagaa tgcagtttag caccgtgttt
      301 tagagctaga aatagcaagt taaaataagg ctagtccgtt atcaacttga aaaagtggca
      361 ccgagtcggt gctttttttt tctttttggc gcgccattaa catgcatgtc ccgcgttcgt
      421 gaaattctct gcatcagcgg agtgatcagg aatcatcgtc tcagcgggat gacgttgcgg
      481 agcaggccgg ctcgcggtgg gcagtcagat gccgaaggcg taactcagga cggcttgcgc
      541 tcatcgcaga acaggggtgg tgcctgcatt gggtgcggtt ggtgatcctg gttggaccgg
      601 tggagatgcg cgcgcacgaa ggggatgtgt cagaaacatt ttgtttgttc tctgtgaact
      661 tttagatgtg ttaaaggcgg cgaatattag cagagagtcc tccttgttgg attctctctt
      721 gaatttcgcc ctttccttct ctttgcgagt ctcgtagaga acaagcactc gttcgccgtc
      781 cctgacgacg caacccgcgc agaagacatc caccaaacgg tgttacacaa tcaccttgtg
      841 tgaagttctt gcggaaaact actcgttggc attttttctt gaattccctt tttcgacaca
      901 attgatggac tataaggacc acgacggaga ctacaaggat catgatattg attacaaaga
      961 cgatgacgat aagatggccc caaagaagaa gcggaaggtc ggtatccacg gagtcccagc
     1021 agccgacaag aagtacagca tcggcctgga catcggcacc aactctgtgg gctgggccgt
     1081 gatcaccgac gagtacaagg tgcccagcaa gaaattcaag gtgctgggca acaccgaccg
     1141 gcacagcatc aagaagaacc tgatcggagc cctgctgttc gacagcggcg aaacagccga
     1201 ggccacccgg ctgaagagaa ccgccagaag aagatacacc agacggaaga accggatctg
     1261 ctatctgcaa gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccacag
     1321 actggaagag tccttcctgg tggaagagga taagaagcac gagcggcacc ccatcttcgg
     1381 caacatcgtg gacgaggtgg cctaccacga gaagtacccc accatctacc acctgagaaa
     1441 gaaactggtg gacagcaccg acaaggccga cctgcggctg atctatctgg ccctggccca
     1501 catgatcaag ttccggggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga
     1561 cgtggacaag ctgttcatcc agctggtgca gacctacaac cagctgttcg aggaaaaccc
     1621 catcaacgcc agcggcgtgg acgccaaggc catcctgtct gccagactga gcaagagcag
     1681 acggctggaa aatctgatcg cccagctgcc cggcgagaag aagaatggcc tgttcggaaa
     1741 cctgattgcc ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga
     1801 ggatgccaaa ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc
     1861 ccagatcggc gaccagtacg ccgacctgtt tctggccgcc aagaacctgt ccgacgccat
     1921 cctgctgagc gacatcctga gagtgaacac cgagatcacc aaggcccccc tgagcgcctc
     1981 tatgatcaag agatacgacg agcaccacca ggacctgacc ctgctgaaag ctctcgtgcg
     2041 gcagcagctg cctgagaagt acaaagagat tttcttcgac cagagcaaga acggctacgc
     2101 cggctacatt gacggcggag ccagccagga agagttctac aagttcatca agcccatcct
     2161 ggaaaagatg gacggcaccg aggaactgct cgtgaagctg aacagagagg acctgctgcg
     2221 gaagcagcgg accttcgaca acggcagcat cccccaccag atccacctgg gagagctgca
     2281 cgccattctg cggcggcagg aagattttta cccattcctg aaggacaacc gggaaaagat
     2341 cgagaagatc ctgaccttcc gcatccccta ctacgtgggc cctctggcca ggggaaacag
     2401 cagattcgcc tggatgacca gaaagagcga ggaaaccatc accccctgga acttcgagga
     2461 agtggtggac aagggcgctt ccgcccagag cttcatcgag cggatgacca acttcgataa
     2521 gaacctgccc aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt
     2581 gtataacgag ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc ccgccttcct
     2641 gagcggcgag cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc ggaaagtgac
     2701 cgtgaagcag ctgaaagagg actacttcaa gaaaatcgag tgcttcgact ccgtggaaat
     2761 ctccggcgtg gaagatcggt tcaacgcctc cctgggcaca taccacgatc tgctgaaaat
     2821 tatcaaggac aaggacttcc tggacaatga ggaaaacgag gacattctgg aagatatcgt
     2881 gctgaccctg acactgtttg aggacagaga gatgatcgag gaacggctga aaacctatgc
     2941 ccacctgttc gacgacaaag tgatgaagca gctgaagcgg cggagataca ccggctgggg
     3001 caggctgagc cggaagctga tcaacggcat ccgggacaag cagtccggca agacaatcct
     3061 ggatttcctg aagtccgacg gcttcgccaa cagaaacttc atgcagctga tccacgacga
     3121 cagcctgacc tttaaagagg acatccagaa agcccaggtg tccggccagg gcgatagcct
     3181 gcacgagcac attgccaatc tggccggcag ccccgccatt aagaagggca tcctgcagac
     3241 agtgaaggtg gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg agaacatcgt
     3301 gatcgaaatg gccagagaga accagaccac ccagaaggga cagaagaaca gccgcgagag
     3361 aatgaagcgg atcgaagagg gcatcaaaga gctgggcagc cagatcctga aagaacaccc
     3421 cgtggaaaac acccagctgc agaacgagaa gctgtacctg tactacctgc agaatgggcg
     3481 ggatatgtac gtggaccagg aactggacat caaccggctg tccgactacg atgtggacca
     3541 tatcgtgcct cagagctttc tgaaggacga ctccatcgac aacaaggtgc tgaccagaag
     3601 cgacaagaac cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga agaagatgaa
     3661 gaactactgg cggcagctgc tgaacgccaa gctgattacc cagagaaagt tcgacaatct
     3721 gaccaaggcc gagagaggcg gcctgagcga actggataag gccggcttca tcaagagaca
     3781 gctggtggaa acccggcaga tcacaaagca cgtggcacag atcctggact cccggatgaa
     3841 cactaagtac gacgagaatg acaagctgat ccgggaagtg aaagtgatca ccctgaagtc
     3901 caagctggtg tccgatttcc ggaaggattt ccagttttac aaagtgcgcg agatcaacaa
     3961 ctaccaccac gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc tgatcaaaaa
     4021 gtaccctaag ctggaaagcg agttcgtgta cggcgactac aaggtgtacg acgtgcggaa
     4081 gatgatcgcc aagagcgagc aggaaatcgg caaggctacc gccaagtact tcttctacag
     4141 caacatcatg aactttttca agaccgagat taccctggcc aacggcgaga tccggaagcg
     4201 gcctctgatc gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg gccgggattt
     4261 tgccaccgtg cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa agaccgaggt
     4321 gcagacaggc ggcttcagca aagagtctat cctgcccaag aggaacagcg ataagctgat
     4381 cgccagaaag aaggactggg accctaagaa gtacggcggc ttcgacagcc ccaccgtggc
     4441 ctattctgtg ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac tgaagagtgt
     4501 gaaagagctg ctggggatca ccatcatgga aagaagcagc ttcgagaaga atcccatcga
     4561 ctttctggaa gccaagggct acaaagaagt gaaaaaggac ctgatcatca agctgcctaa
     4621 gtactccctg ttcgagctgg aaaacggccg gaagagaatg ctggcctctg ccggcgaact
     4681 gcagaaggga aacgaactgg ccctgccctc caaatatgtg aacttcctgt acctggccag
     4741 ccactatgag aagctgaagg gctcccccga ggataatgag cagaaacagc tgtttgtgga
     4801 acagcacaag cactacctgg acgagatcat cgagcagatc agcgagttct ccaagagagt
     4861 gatcctggcc gacgctaatc tggacaaagt gctgtccgcc tacaacaagc accgggataa
     4921 gcccatcaga gagcaggccg agaatatcat ccacctgttt accctgacca atctgggagc
     4981 ccctgccgcc ttcaagtact ttgacaccac catcgaccgg aagaggtaca ccagcaccaa
     5041 agaggtgctg gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacacggat
     5101 cgacctgtct cagctgggag gcgacaaaag gccggcggcc acgaaaaagg ccggccaggc
     5161 aaaaaagaaa aagcatatgg gaagcggagc tactaacttc agcctgctga agcaggctgg
     5221 agacgtggag gagaaccctg gacctgctag cggaatgcat gagaaaaaaa tcactggata
     5281 taccaccgtt gatatatccc aatggcatcg taaagaacat tttgaggcat ttcagtcagt
     5341 tgctcaatgt acctataacc agaccgttca gctggatatt acggcctttt taaagaccgt
     5401 aaagaaaaat aagcacaagt tttatccggc atttattcac attcttgccc gcctgatgaa
     5461 tgctcatccg gaattccgta tggcaatgaa agacggtgag ctggtgatat gggatagtgt
     5521 tcacccttgt tacaccgttt tccatgagca aactgaaacg ttttcatcgc tctggagtga
     5581 ataccacgac gatttccggc agtttctaca catatattcg caagatgtgg cgtgttacgg
     5641 tgaaaacctg gcctatttcc ctaaagggtt tattgagaat atgtttttcg tctcagccaa
     5701 tccctgggtg agtttcacca gttttgattt aaacgtggcc aatatggaca acttcttcgc
     5761 ccccgttttc accatgggca aatattatac gcaaggcgac aaggtgctga tgccgctggc
     5821 gattcaggtt catcatgccg tttgtgatgg cttccatgtc ggcagaatgc ttaatgaatt
     5881 acaacagtac tgcgatgagt ggcagggcgg ggcttaccca tacgatgttc ctgactatgc
     5941 gtaaggatcc acagaagctg cccgtctctc gttttcctct cttttcggag ggatcaggga
     6001 gagtgcctcg ggtcggagag agctgacgag ggggtgccag agacccctgt gtcctttatc
     6061 gaagaaaagg gatgactctt catgtggcat ttcacacagt ctcacctcgc cttgttttct
     6121 ttttgtcaat cagaacgaaa gcgagttgcg ggtgacgcag atgtgcgtgt atccactcgt
     6181 gaatgcgtta tcgttctgta tgccgctaga gtgctggact gttgctgtct gcccacgaca
     6241 gcagacaact ttccttctat gcacttgcag gatggtgcag cgcaaacgac ggagagaaag
     6301 gagcaccctc tcagtttccc tacgatgtgc tgtcagtttc gactcttcac cgcgaacgat
     6361 tggcgatacg tctctgttga cttgttaggc tccgaccacg aagctccctt aactagataa
     6421 gccgcgacac ctaagtgtac accatttgca gatcgataat ctgcgaccgc tgaatccgtc
     6481 cagatcagta aaaccgcacc acctaagtgt aaaccttgtt taggtcgata aaatgctacc
     6541 aacccccacc cacaatcgag ccttgagcgt ttctgcgcac gcgttggcct acgtgacttg
     6601 ctgatgcctg cctctggcca ttcatgccag tcagtgcgca taaaaatgtg gacacagtcg
     6661 gttgacaagt gttctggcag gctacagtga cacgcggccg cagctccagc ttttgttccc
     6721 tttagtgagg gttaatttcg agcttggcgt aatcatggtc atagctgttt cctgtgtgaa
     6781 attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct
     6841 ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc
     6901 agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg
     6961 gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc
     7021 ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag
     7081 gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa
     7141 aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc
     7201 gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     7261 ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg
     7321 cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt
     7381 cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc
     7441 gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc
     7501 cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag
     7561 agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg
     7621 ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa
     7681 ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag
     7741 gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact
     7801 cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa
     7861 attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt
     7921 accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag
     7981 ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca
     8041 gtgctgcaat gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc
     8101 agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt
     8161 ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg
     8221 ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca
     8281 gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg
     8341 ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca
     8401 tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg
     8461 tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct
     8521 cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca
     8581 tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca
     8641 gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg
     8701 tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac
     8761 ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt
     8821 attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc
     8881 cgcgcacatt tccccgaaaa gtgccaccta aattgtaagc gttaatattt tgttaaaatt
     8941 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa taggccgaaa tcggcaaaat
     9001 cccttataaa tcaaaagaat agaccgagat agggttgagt gttgttccag tttggaacaa
     9061 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     9121 cgatggccca ctacgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa
     9181 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     9241 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag
     9301 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     9361 cgcgtcccat tcgccattca ggctgcgcaa ctgttgggaa gggcgatcgg tgcgggcctc
     9421 ttcgctatta cgccagctgg cgaaaggggg atgtgctgca aggcgattaa gttgggtaac
     9481 gccagggttt tcccagtcac gacgttgtaa aacgacggcc agtgaattgt aatacgactc
     9541 actatagggc gaattgggta ccgggccccc cctcgaggtc gacggtatcg atattaatta
     9601 agatgagaca aagtgcgcga gttgaaatcg tcgtggggac gatttcaccg cggccacatg
     9661 ttggagacac tgagggcaca cgggaaacgc gaaagatttc aaattaacgt acccaaacgc
     9721 gaaagcttgc gcagcataca ctcgaagcga acatcccgaa ccatcgagag gcggagagcg
     9781 ataagtcttt cacgctgcga agtgttgcga cggctgcgcc gctgca
//
