LOCUS       sequence-313847-       10641 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  inducible expression of Cas9.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10641)
  TITLE     Lian Lab PB plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 10641)
  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: 
            "Unpublished"
COMMENT     Sequence Label: PB-iCas9
FEATURES             Location/Qualifiers
     source          1..10641
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      21..30
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             30..95
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             102..122
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             147..4247
                     /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             147..4247
                     /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             4248..4295
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     promoter        4305..4815
                     /label=hPGK promoter
                     /note="human phosphoglycerate kinase 1 promoter"
     primer_bind     4734..4752
                     /label=hPGK-F
                     /note="Human PGK promoter, forward primer"
     CDS             4825..5421
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4825..4844)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5321..5341
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     CDS             5422..5475
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(5476..6093,6094..6099,6100..6135,6136..6138,6139..6174,
                     6175..6177,6178..6213,6214..6222)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     primer_bind     complement(6262..6282)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6680..6691)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     repeat_region   6979..7041
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7092..7112)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(7092..7110)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(7131..7147)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7131..7147)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7144..7166)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7155..7171
                     /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(7179..7185,7186..7203,7204..7209))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7224..7245
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7362..7379)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7533..8121)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7613..7632)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8292..9083,9084..9152))
                     /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     8915..8934
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9153..9257)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(9283..9738)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9420..9441)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     9632..9651
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9865..9887
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     9879..9896
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     9880..9896
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9906..9925
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        9906..9924
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   9954..9988
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        join(10331..10641,1..4)
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    10339..10357
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10375..10393
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10410..10428
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10446..10464
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10482..10500
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10517..10535
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    10553..10571
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     join(10618..10641,1)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
ORIGIN
        1 cgcctggaga attggctagc gccaccatgg actataagga ccacgacgga gactacaagg
       61 atcatgatat tgattacaaa gacgatgacg ataagatggc cccaaagaag aagcggaagg
      121 tcggtatcca cggagtccca gcagccgaca agaagtacag catcggcctg gacatcggca
      181 ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca
      241 aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt
      301 tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca
      361 ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg
      421 tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc
      481 acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc
      541 ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc
      601 tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg
      661 acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca
      721 accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt
      781 ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga
      841 agaagaatgg cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca
      901 agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg
      961 acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg
     1021 ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca
     1081 ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga
     1141 ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg
     1201 accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct
     1261 acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc
     1321 tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc
     1381 agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc
     1441 tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg
     1501 gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca
     1561 tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg
     1621 agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc
     1681 tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg
     1741 gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt
     1801 tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg
     1861 agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca
     1921 cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg
     1981 aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg
     2041 aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc
     2101 ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca
     2161 agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact
     2221 tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg
     2281 tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca
     2341 ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc
     2401 ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg
     2461 gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca
     2521 gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc
     2581 tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc
     2641 tgtccgacta cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg
     2701 acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg
     2761 aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta
     2821 cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata
     2881 aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac
     2941 agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag
     3001 tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt
     3061 acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg
     3121 tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact
     3181 acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta
     3241 ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg
     3301 ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg
     3361 tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga
     3421 atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca
     3481 agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg
     3541 gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca
     3601 agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca
     3661 gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg
     3721 acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa
     3781 tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg
     3841 tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg
     3901 agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga
     3961 tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg
     4021 cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt
     4081 ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc
     4141 ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca
     4201 ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaaa aggccggcgg
     4261 ccacgaaaaa ggccggccag gcaaaaaaga aaaagtaagg atccggggtt ggggttgcgc
     4321 cttttccaag gcagccctgg gtttgcgcag ggacgcggct gctctgggcg tggttccggg
     4381 aaacgcagcg gcgccgaccc tgggtctcgc acattcttca cgtccgttcg cagcgtcacc
     4441 cggatcttcg ccgctaccct tgtgggcccc ccggcgacgc ttcctgctcc gcccctaagt
     4501 cgggaaggtt ccttgcggtt cgcggcgtgc cggacgtgac aaacggaagc cgcacgtctc
     4561 actagtaccc tcgcagacgg acagcgccag ggagcaatgg cagcgcgccg accgcgatgg
     4621 gctgtggcca atagcggctg ctcagcaggg cgcgccgaga gcagcggccg ggaaggggcg
     4681 gtgcgggagg cggggtgtgg ggcggtagtg tgggccctgt tcctgcccgc gcggtgttcc
     4741 gcattctgca agcctccgga gcgcacgtcg gcagtcggct ccctcgttga ccgaatcacc
     4801 gacctctctc cccagcaatt caccatgacc gagtacaagc ccacggtgcg cctcgccacc
     4861 cgcgacgacg tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc
     4921 acgcgccaca ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc
     4981 ttcctcacgc gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg
     5041 gtggcggtct ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc
     5101 ccgcgcatgg ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc
     5161 ctggcgccgc accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg cgtctcgccc
     5221 gaccaccagg gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag
     5281 cgcgccgggg tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag
     5341 cggctcggct tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc
     5401 atgacccgca agcccggtgc cgaaggtaga ggttctctcc tcacttgtgg tgatgttgaa
     5461 gaaaaccctg gtccaatgtc tagactggac aagagcaaag tcataaacgg agctctggaa
     5521 ttactcaatg gtgtcggtat cgaaggcctg acgacaagga aactcgctca aaagctggga
     5581 gttgagcagc ctaccctgta ctggcacgtg aagaacaagc gggccctgct cgatgccctg
     5641 ccaatcgaga tgctggacag gcatcatacc cacttctgcc ccctggaagg cgagtcatgg
     5701 caagactttc tgcggaacaa cgccaagtca taccgctgtg ctctcctctc acatcgcgac
     5761 ggggctaaag tgcatctcgg cacccgccca acagagaaac agtacgaaac cctggaaaat
     5821 cagctcgcgt tcctgtgtca gcaaggcttc tccctggaga acgcactgta cgctctgtcc
     5881 gccgtgggcc actttacact gggctgcgta ttggaggaac aggagcatca agtagcaaaa
     5941 gaggaaagag agacacctac caccgattct atgcccccac ttctgagaca agcaattgag
     6001 ctgttcgacc ggcagggagc cgaacctgcc ttccttttcg gcctggaact aatcatatgt
     6061 ggcctggaga aacagctaaa gtgcgaaagc ggcgggccga ccgacgccct tgacgatttt
     6121 gacttagaca tgctcccagc cgatgccctt gacgactttg accttgatat gctgcctgct
     6181 gacgctcttg acgattttga ccttgacatg ctccccgggt aactaagtaa ggatcgatcc
     6241 aagatatcgt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     6301 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     6361 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     6421 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     6481 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     6541 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     6601 gaagctgacg tcctttccat ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac
     6661 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     6721 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     6781 ttgggccgcc tccccgcact agttaaaagt tttgttactt tatagaagaa attttgagtt
     6841 tttgtttttt tttaataaat aaataaacat aaataaattg tttgttgaat ttattattag
     6901 tatgtaagtg taaatataat aaaacttaat atctattcaa attaataaat aaacctcgat
     6961 atacagaccg ataaaacaca tgcgtcaatt ttacgcatga ttatctttaa cgtacgtcac
     7021 aatatgatta tctttctagg gttaatctag tatacgcgta tgcggccgct taattaatcc
     7081 agcttttgtt ccctttagtg agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg
     7141 tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc cggaagcata
     7201 aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc gttgcgctca
     7261 ctgcccgctt tccagtcggg aaacctgtcg tgccagctgc attaatgaat cggccaacgc
     7321 gcggggagag gcggtttgcg tattgggcgc tcttccgctt cctcgctcac tgactcgctg
     7381 cgctcggtcg ttcggctgcg gcgagcggta tcagctcact caaaggcggt aatacggtta
     7441 tccacagaat caggggataa cgcaggaaag aacatgtgag caaaaggcca gcaaaaggcc
     7501 aggaaccgta aaaaggccgc gttgctggcg tttttccata ggctccgccc ccctgacgag
     7561 catcacaaaa atcgacgctc aagtcagagg tggcgaaacc cgacaggact ataaagatac
     7621 caggcgtttc cccctggaag ctccctcgtg cgctctcctg ttccgaccct gccgcttacc
     7681 ggatacctgt ccgcctttct cccttcggga agcgtggcgc tttctcatag ctcacgctgt
     7741 aggtatctca gttcggtgta ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc
     7801 gttcagcccg accgctgcgc cttatccggt aactatcgtc ttgagtccaa cccggtaaga
     7861 cacgacttat cgccactggc agcagccact ggtaacagga ttagcagagc gaggtatgta
     7921 ggcggtgcta cagagttctt gaagtggtgg cctaactacg gctacactag aagaacagta
     7981 tttggtatct gcgctctgct gaagccagtt accttcggaa aaagagttgg tagctcttga
     8041 tccggcaaac aaaccaccgc tggtagcggt ggtttttttg tttgcaagca gcagattacg
     8101 cgcagaaaaa aaggatctca agaagatcct ttgatctttt ctacggggtc tgacgctcag
     8161 tggaacgaaa actcacgtta agggattttg gtcatgagat tatcaaaaag gatcttcacc
     8221 tagatccttt taaattaaaa atgaagtttt aaatcaatct aaagtatata tgagtaaact
     8281 tggtctgaca gttaccaatg cttaatcagt gaggcaccta tctcagcgat ctgtctattt
     8341 cgttcatcca tagttgcctg actccccgtc gtgtagataa ctacgatacg ggagggctta
     8401 ccatctggcc ccagtgctgc aatgataccg cgagacccac gctcaccggc tccagattta
     8461 tcagcaataa accagccagc cggaagggcc gagcgcagaa gtggtcctgc aactttatcc
     8521 gcctccatcc agtctattaa ttgttgccgg gaagctagag taagtagttc gccagttaat
     8581 agtttgcgca acgttgttgc cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt
     8641 atggcttcat tcagctccgg ttcccaacga tcaaggcgag ttacatgatc ccccatgttg
     8701 tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg tcagaagtaa gttggccgca
     8761 gtgttatcac tcatggttat ggcagcactg cataattctc ttactgtcat gccatccgta
     8821 agatgctttt ctgtgactgg tgagtactca accaagtcat tctgagaata gtgtatgcgg
     8881 cgaccgagtt gctcttgccc ggcgtcaata cgggataata ccgcgccaca tagcagaact
     8941 ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa aactctcaag gatcttaccg
     9001 ctgttgagat ccagttcgat gtaacccact cgtgcaccca actgatcttc agcatctttt
     9061 actttcacca gcgtttctgg gtgagcaaaa acaggaaggc aaaatgccgc aaaaaaggga
     9121 ataagggcga cacggaaatg ttgaatactc atactcttcc tttttcaata ttattgaagc
     9181 atttatcagg gttattgtct catgagcgga tacatatttg aatgtattta gaaaaataaa
     9241 caaatagggg ttccgcgcac atttccccga aaagtgccac ctaaattgta agcgttaata
     9301 ttttgttaaa attcgcgtta aatttttgtt aaatcagctc attttttaac caataggccg
     9361 aaatcggcaa aatcccttat aaatcaaaag aatagaccga gatagggttg agtgttgttc
     9421 cagtttggaa caagagtcca ctattaaaga acgtggactc caacgtcaaa gggcgaaaaa
     9481 ccgtctatca gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt
     9541 cgaggtgccg taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac
     9601 ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta
     9661 gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg
     9721 cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg caactgttgg gaagggcgat
     9781 cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat
     9841 taagttgggt aacgccaggg ttttcccagt cacgacgttg taaaacgacg gccagtgagc
     9901 gcgcgtaata cgactcacta tagggcgaat tggggcgcgc cattctagat taaccctaga
     9961 aagatagtct gcgtaaaatt gacgcatgca ttcttgaaat attgctctct ctttctaaat
    10021 agcgcgaatc cgtcgctgtg catttaggac atctcagtcg ccgcttggag ctcccgtgag
    10081 gcgtgcttgt caatgcggta agtgtcactg attttgaact ataacgaccg cgtgagtcaa
    10141 aatgacgcat gattatcttt tacgtgactt ttaagattta actcatacga taattatatt
    10201 gttatttcat gttctactta cgtgataact tattatatat atattttctt gttatagata
    10261 tcaactagaa tgctagccgg gtttattaca gggacagcag agatccactt tggccgcgaa
    10321 tcgatatgtc gagtttactc cctatcagtg atagagaacg tatgtcgagt ttactcccta
    10381 tcagtgatag agaacgatgt cgagtttact ccctatcagt gatagagaac gtatgtcgag
    10441 tttactccct atcagtgata gagaacgtat gtcgagttta ctccctatca gtgatagaga
    10501 acgtatgtcg agtttatccc tatcagtgat agagaacgta tgtcgagttt actccctatc
    10561 agtgatagag aacgtatgtc gaggtaggcg tgtacggtgg gaggcctata taagcagagc
    10621 tcgtttagtg aaccgtcaga t
//
