LOCUS       sequence-320739-       16256 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CRISPR plasmid with spCas9 expressed under egg cell promoter - 
            TagRFP seed selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 16256)
  TITLE     CRISPR plasmid (spCas9 under EC1.2/EC1.1 promoter)
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 16256)
  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            "Unpublished"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: pEC1.2/EC1.1_spCas9_TagRFP
FEATURES             Location/Qualifiers
     source          1..16256
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1362..1427
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1434..1454
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1479..5579
                     /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             1479..5579
                     /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"
     RBS             5492..5500
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             5580..5627
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6842..7537
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFMYGSRTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEANTEMLYPADGGLEGRSDMALK
                     LVGGGHLICNFKTTYRSKKPAKNLKMPGVYYVDHRLERIKEADKETYVEQHEVAVARYC
                     DLPSKLGHK"
     terminator      complement(7536..7787)
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     misc_feature    7794..7818
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     CDS             8243..9037
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      9124..9712
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9613..9632
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9866..9883
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    9898..10038
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(9964..9983)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     10124..10146
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     rep_origin      10382..10576
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     CDS             complement(10642..11715)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     CDS             complement(12144..12773)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     misc_feature    14074..14098
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     primer_bind     14286..14308
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     14300..14317
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     14301..14317
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             15030..15821
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     misc_RNA        15973..16048
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 gaataaaagc atttgcgttt ggtttatcat tgcgtttata caaggacaga gatccactga
       61 gctggaatag cttaaaacca ttatcagaac aaaataaacc attttttgtt aagaatcaga
      121 gcatagtaaa caacagaaac aacctaagag aggtaacttg tccaagaaga tagctaatta
      181 tatctatttt ataaaagtta tcatagtttg taagtcacaa aagatgcaaa taacagagaa
      241 actaggagac ttgagaatat acattcttgt atatttgtat tcgagattgt gaaaatttga
      301 ccataagttt aaattcttaa aaagatatat ctgatctagg tgatggttat agactgtaat
      361 tttaccacat gtttaatgat ggatagtgac acacatgaca catcgacaac actatagcat
      421 cttatttaga ttacaacatg aaatttttct gtaatacatg tctttgtaca taatttaaaa
      481 gtaattccta agaaatatat ttatacaagg agtttaaaga aaacatagca taaagttcaa
      541 tgagtagtaa aaaccatata cagtatatag cataaagttc aatgagttta ttacaaaagc
      601 attggttcac tttctgtaac acgacgttaa accttcgtct ccaataggag cgctactgat
      661 tcaacatgcc aatatatact aaatacgttt ctacagtcaa atgctttaac gtttcatgat
      721 taagtgacta tttaccgtca atcctttccc attcctccca ctaatccaac tttttaatta
      781 ctcttaaatc accactaagc tagtaacgcc tatcatgaat tagctctact aaatctagca
      841 acctttcaaa tttgcagtat tgcaggtgtc tctgtgtctt taaaatagtt gccttatgat
      901 ttcttcggtt tcaagatgat caaatagtta tagatttcat gctcacacat gctcattaga
      961 tgtgtacata ctttacttac ccaaatctat tttctcgcaa agattttgat ggtaaagctg
     1021 atttggttct attgaactaa atcaaacgag tttcagactg agtgattcta atccggccca
     1081 ttagccccta aacagaccca ctaattacgc agcttttaat agagtaatta cacctagttt
     1141 acccactaaa ccactaagca ctaattatct cacaatctaa tgagcttccc tcgtaattac
     1201 ttgggctttc actctaccat ttatttgtaa cagtcaagtc tctactgtct ctatataaac
     1261 tctctaaagt taacacacaa ttctcatcac aaacaaatca accaaagcaa cttctactct
     1321 ttcttctttc gaccttatca atctgttgag aaatctagat ggattacaag gaccacgacg
     1381 gggattacaa ggaccacgac attgattaca aggatgatga tgacaagatg gctccgaaga
     1441 agaagaggaa ggttggcatc cacggggtgc cagctgctga caagaagtac tcgatcggcc
     1501 tcgatattgg gactaactct gttggctggg ccgtgatcac cgacgagtac aaggtgccct
     1561 caaagaagtt caaggtcctg ggcaacaccg atcggcattc catcaagaag aatctcattg
     1621 gcgctctcct gttcgacagc ggcgagacgg ctgaggctac gcggctcaag cgcaccgccc
     1681 gcaggcggta cacgcgcagg aagaatcgca tctgctacct gcaggagatt ttctccaacg
     1741 agatggcgaa ggttgacgat tctttcttcc acaggctgga ggagtcattc ctcgtggagg
     1801 aggataagaa gcacgagcgg catccaatct tcggcaacat tgtcgacgag gttgcctacc
     1861 acgagaagta ccctacgatc taccatctgc ggaagaagct cgtggactcc acagataagg
     1921 cggacctccg cctgatctac ctcgctctgg cccacatgat taagttcagg ggccatttcc
     1981 tgatcgaggg ggatctcaac ccggacaata gcgatgttga caagctgttc atccagctcg
     2041 tgcagacgta caaccagctc ttcgaggaga accccattaa tgcgtcaggc gtcgacgcga
     2101 aggctatcct gtccgctagg ctctcgaagt ctcggcgcct cgagaacctg atcgcccagc
     2161 tgccgggcga gaagaagaac ggcctgttcg ggaatctcat tgcgctcagc ctggggctca
     2221 cgcccaactt caagtcgaat ttcgatctcg ctgaggacgc caagctgcag ctctccaagg
     2281 acacatacga cgatgacctg gataacctcc tggcccagat cggcgatcag tacgcggacc
     2341 tgttcctcgc tgccaagaat ctgtcggacg ccatcctcct gtctgatatt ctcagggtga
     2401 acaccgagat tacgaaggct ccgctctcag cctccatgat caagcgctac gacgagcacc
     2461 atcaggatct gaccctcctg aaggcgctgg tcaggcagca gctccccgag aagtacaagg
     2521 agatcttctt cgatcagtcg aagaacggct acgctgggta cattgacggc ggggcctctc
     2581 aggaggagtt ctacaagttc atcaagccga ttctggagaa gatggacggc acggaggagc
     2641 tgctggtgaa gctcaatcgc gaggacctcc tgaggaagca gcggacattc gataacggca
     2701 gcatcccaca ccagattcat ctcggggagc tgcacgctat cctgaggagg caggaggact
     2761 tctacccttt cctcaaggat aaccgcgaga agatcgagaa gattctgact ttcaggatcc
     2821 cgtactacgt cggcccactc gctaggggca actcccgctt cgcttggatg acccgcaagt
     2881 cagaggagac gatcacgccg tggaacttcg aggaggtggt cgacaagggc gctagcgctc
     2941 agtcgttcat cgagaggatg acgaatttcg acaagaacct gccaaatgag aaggtgctcc
     3001 ctaagcactc gctcctgtac gagtacttca cagtctacaa cgagctgact aaggtgaagt
     3061 atgtgaccga gggcatgagg aagccggctt tcctgtctgg ggagcagaag aaggccatcg
     3121 tggacctcct gttcaagacc aaccggaagg tcacggttaa gcagctcaag gaggactact
     3181 tcaagaagat tgagtgcttc gattcggtcg agatctctgg cgttgaggac cgcttcaacg
     3241 cctccctggg gacctaccac gatctcctga agatcattaa ggataaggac ttcctggaca
     3301 acgaggagaa tgaggatatc ctcgaggaca ttgtgctgac actcactctg ttcgaggacc
     3361 gggagatgat cgaggagcgc ctgaagactt acgcccatct cttcgatgac aaggtcatga
     3421 agcagctcaa gaggaggagg tacaccggct gggggaggct gagcaggaag ctcatcaacg
     3481 gcattcggga caagcagtcc gggaagacga tcctcgactt cctgaagagc gatggcttcg
     3541 cgaaccgcaa tttcatgcag ctgattcacg atgacagcct cacattcaag gaggatatcc
     3601 agaaggctca ggtgagcggc cagggggact cgctgcacga gcatatcgcg aacctcgctg
     3661 gctcgccagc tatcaagaag gggattctgc agaccgtgaa ggttgtggac gagctggtga
     3721 aggtcatggg caggcacaag cctgagaaca tcgtcattga gatggcccgg gagaatcaga
     3781 ccacgcagaa gggccagaag aactcacgcg agaggatgaa gaggatcgag gagggcatta
     3841 aggagctggg gtcccagatc ctcaaggagc acccggtgga gaacacgcag ctgcagaatg
     3901 agaagctcta cctgtactac ctccagaatg gccgcgatat gtatgtggac caggagctgg
     3961 atattaacag gctcagcgat tacgacgtcg atcatatcgt tccacagtca ttcctgaagg
     4021 atgactccat tgacaacaag gtcctcacca ggtcggacaa gaaccggggc aagtctgata
     4081 atgttccttc agaggaggtc gttaagaaga tgaagaacta ctggcgccag ctcctgaatg
     4141 ccaagctgat cacgcagcgg aagttcgata acctcacaaa ggctgagagg ggcgggctct
     4201 ctgagctgga caaggcgggc ttcatcaaga ggcagctggt cgagacacgg cagatcacta
     4261 agcacgttgc gcagattctc gactcacgga tgaacactaa gtacgatgag aatgacaagc
     4321 tgatccgcga ggtgaaggtc atcaccctga agtcaaagct cgtctccgac ttcaggaagg
     4381 atttccagtt ctacaaggtt cgggagatca acaattacca ccatgcccat gacgcgtacc
     4441 tgaacgcggt ggtcggcaca gctctgatca agaagtaccc aaagctcgag agcgagttcg
     4501 tgtacgggga ctacaaggtt tacgatgtga ggaagatgat cgccaagtcg gagcaggaga
     4561 ttggcaaggc taccgccaag tacttcttct actctaacat tatgaatttc ttcaagacag
     4621 agatcactct ggccaatggc gagatccgga agcgccccct catcgagacg aacggcgaga
     4681 cgggggagat cgtgtgggac aagggcaggg atttcgcgac cgtcaggaag gttctctcca
     4741 tgccacaagt gaatatcgtc aagaagacag aggtccagac tggcgggttc tctaaggagt
     4801 caattctgcc taagcggaac agcgacaagc tcatcgcccg caagaaggac tgggatccga
     4861 agaagtacgg cgggttcgac agccccactg tggcctactc ggtcctggtt gtggcgaagg
     4921 ttgagaaggg caagtccaag aagctcaaga gcgtgaagga gctgctgggg atcacgatta
     4981 tggagcgctc cagcttcgag aagaacccga tcgatttcct ggaggcgaag ggctacaagg
     5041 aggtgaagaa ggacctgatc attaagctcc ccaagtactc actcttcgag ctggagaacg
     5101 gcaggaagcg gatgctggct tccgctggcg agctgcagaa ggggaacgag ctggctctgc
     5161 cgtccaagta tgtgaacttc ctctacctgg cctcccacta cgagaagctc aagggcagcc
     5221 ccgaggacaa cgagcagaag cagctgttcg tcgagcagca caagcattac ctcgacgaga
     5281 tcattgagca gatttccgag ttctccaagc gcgtgatcct ggccgacgcg aatctggata
     5341 aggtcctctc cgcgtacaac aagcaccgcg acaagccaat cagggagcag gctgagaata
     5401 tcattcatct cttcaccctg acgaacctcg gcgcccctgc tgctttcaag tacttcgaca
     5461 caactatcga tcgcaagagg tacacaagca ctaaggaggt cctggacgcg accctcatcc
     5521 accagtcgat taccggcctc tacgagacgc gcatcgacct gtctcagctc gggggcgaca
     5581 agcggccagc ggcgacgaag aaggcggggc aggcgaagaa gaagaagtga gctcagagct
     5641 ttcgttcgta tcatcggttt cgacaacgtt cgtcaagttc aatgcatcag tttcattgcg
     5701 cacacaccag aatcctactg agtttgagta ttatggcatt gggaaaactg tttttcttgt
     5761 accatttgtt gtgcttgtaa tttactgtgt tttttattcg gttttcgcta tcgaactgtg
     5821 aaatggaaat ggatggagaa gagttaatga atgatatggt ccttttgttc attctcaaat
     5881 taatattatt tgttttttct cttatttgtt gtgtgttgaa tttgaaatta taagagatat
     5941 gcaaacattt tgttttgagt aaaaatgtgt caaatcgtgg cctctaatga ccgaagttaa
     6001 tatgaggagt aaaacacttg tagttgtacc attatgctta ttcactaggc aacaaatata
     6061 ttttcagacc tagaaaagct gcaaatgtta ctgaatacaa gtatgtcctc ttgtgtttta
     6121 gacatttatg aactttcctt tatgtaattt tccagaatcc ttgtcagatt ctaatcattg
     6181 ctttataatt atagttatac tcatggattt gtagttgagt atgaaaatat tttttaatgc
     6241 attttatgac ttgccaattg attgacaacg aattggtacc gaattggaaa ccaaattaac
     6301 atagggtttt tatttaaata aaagtttaac cttcttttta aaaaattgtt catagtgtca
     6361 tgtcagaaca agagctacaa atcacacata gcatgcataa gcggagctat gatgagtggt
     6421 attgttttgt tcgtcacttg tcactctttt ccaacacata atcccgacaa caacgtaaga
     6481 gcatctctct ctctccacac acactcatgc atgcatgcat tcttacacgt gattgccatg
     6541 caaatctcct ttctcaccta taaatacaaa ccaacccttc actacactct tcactcaaac
     6601 caaaacaaga aaacatacac aaatagcaaa acaagggcga attctcaaca caacatatac
     6661 aaaacaaacg aatctcaagc aatcaagcat tctacttcta ttgcagcaat ttaaatcatt
     6721 tcttttaaag caaaagctat tttctgaaaa ttttcaccat ttacgaacga tagccatggt
     6781 gaagactaat ctttttctct ttttcatctt ttcacttctc ctatcattat cctcggccga
     6841 catgagcgag ctgattaagg agaacatgca catgaagctg tacatggagg gcaccgtgaa
     6901 caaccaccac ttcaagtgca catccgaggg cgaaggcaag ccctacgagg gcacccagac
     6961 catgagaatc aaggtggtcg agggcggccc tctccccttc gccttcgaca tcctggctac
     7021 cagcttcatg tacggcagca gaaccttcat caaccacacc cagggcatcc ccgacttctt
     7081 taagcagtcc ttccctgagg gcttcacatg ggagagagtc accacatacg aagacggggg
     7141 cgtgctgacc gctacccagg acaccagcct ccaggacggc tgcctcatct acaacgtcaa
     7201 gatcagaggg gtgaacttcc catccaacgg ccctgtgatg cagaagaaaa cactcggctg
     7261 ggaggccaac accgagatgc tgtaccccgc tgacggcggc ctggaaggca gaagcgacat
     7321 ggccctgaag ctcgtgggcg ggggccacct gatctgcaac ttcaagacca catacagatc
     7381 caagaaaccc gctaagaacc tcaagatgcc cggcgtctac tatgtggacc acagactgga
     7441 aagaatcaag gaggccgaca aagaaaccta cgtcgagcag cacgaggtgg ctgtggccag
     7501 atactgcgac ctccctagca aactggggca caagtgatct agtaacatag atgacaccgc
     7561 gcgcgataat ttatcctagt ttgcgcgcta tattttgttt tctatcgcgt attaaatgta
     7621 taattgcggg actctaatca taaaaaccca tctcataaat aacgtcatgc attacatgtt
     7681 aattattaca tgcttaacgt aattcaacag aaattatatg ataatcatcg caagaccggc
     7741 aacaggattc aatcttaaga aactttattg ccaaatgttt gaacgatggt accgtttaca
     7801 ccacaatata tcctgccacc agccagccaa cagctccccg accggcagct cggcacaaaa
     7861 tcaccactcg atacaggcag cccatcagtc cgggacggcg tcagcgggag agccgttgta
     7921 aggcggcaga ctttgctcat gttaccgatg ctattcggaa gaacggcaac taagctgccg
     7981 ggtttgaaac acggatgatc tcgcggaggg tagcatgttg attgtaacga tgacagagcg
     8041 ttgctgcctg tgatcaccgc ggtttcaaaa tcggctccgt cgatactatg ttatacgcca
     8101 actttgaaaa caactttgaa aaagctgttt tctggtattt aaggttttag aatgcaagga
     8161 acagtgaatt ggagttcgtc ttgttataat tagcttcttg gggtatcttt aaatactgta
     8221 gaaaagagga aggaaataat aaatggctaa aatgagaata tcaccggaat tgaaaaaact
     8281 gatcgaaaaa taccgctgcg taaaagatac ggaaggaatg tctcctgcta aggtatataa
     8341 gctggtggga gaaaatgaaa acctatattt aaaaatgacg gacagccggt ataaagggac
     8401 cacctatgat gtggaacggg aaaaggacat gatgctatgg ctggaaggaa agctgcctgt
     8461 tccaaaggtc ctgcactttg aacggcatga tggctggagc aatctgctca tgagtgaggc
     8521 cgatggcgtc ctttgctcgg aagagtatga agatgaacaa agccctgaaa agattatcga
     8581 gctgtatgcg gagtgcatca ggctctttca ctccatcgac atatcggatt gtccctatac
     8641 gaatagctta gacagccgct tagccgaatt ggattactta ctgaataacg atctggccga
     8701 tgtggattgc gaaaactggg aagaagacac tccatttaaa gatccgcgcg agctgtatga
     8761 ttttttaaag acggaaaagc ccgaagagga acttgtcttt tcccacggcg acctgggaga
     8821 cagcaacatc tttgtgaaag atggcaaagt aagtggcttt attgatcttg ggagaagcgg
     8881 cagggcggac aagtggtatg acattgcctt ctgcgtccgg tcgatcaggg aggatatcgg
     8941 ggaagaacag tatgtcgagc tattttttga cttactgggg atcaagcctg attgggagaa
     9001 aataaaatat tatattttac tggatgaatt gttttagtac ctagaatgca tgaccaaaat
     9061 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     9121 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     9181 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     9241 cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca
     9301 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     9361 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     9421 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     9481 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     9541 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     9601 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     9661 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     9721 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     9781 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     9841 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgcct
     9901 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct
     9961 cagtacaatc tgctctgatg ccgcatagtt aagccagtat acactccgct atcgctacgt
    10021 gactgggtca tggctgcgcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
    10081 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
    10141 cagaggtttt caccgtcatc accgaaacgc gcgaggcagg gtgccttgat gtgggcgccg
    10201 gcggtcgagt ggcgacggcg cggcttgtcc gcgccctggt agattgcctg gccgtaggcc
    10261 agccattttt gagcggccag cggccgcgat aggccgacgc gaagcggcgg ggcgtaggga
    10321 gcgcagcgac cgaagggtag gcgctttttg cagctcttcg gctgtgcgct ggccagacag
    10381 ttatgcacag gccaggcggg ttttaagagt tttaataagt tttaaagagt tttaggcgga
    10441 aaaatcgcct tttttctctt ttatatcagt cacttacatg tgtgaccggt tcccaatgta
    10501 cggctttggg ttcccaatgt acgggttccg gttcccaatg tacggctttg ggttcccaat
    10561 gtacgtgcta tccacaggaa acagaccttt tcgacctttt tcccctgcta gggcaatttg
    10621 ccctagcatc tgctccgtac attaggaacc ggcggatgct tcgccctcga tcaggttgcg
    10681 gtagcgcatg actaggatcg ggccagcctg ccccgcctcc tccttcaaat cgtactccgg
    10741 caggtcattt gacccgatca gcttgcgcac ggtgaaacag aacttcttga actctccggc
    10801 gctgccactg cgttcgtaga tcgtcttgaa caaccatctg gcttctgcct tgcctgcggc
    10861 gcggcgtgcc aggcggtaga gaaaacggcc gatgccggga tcgatcaaaa agtaatcggg
    10921 gtgaaccgtc agcacgtccg ggttcttgcc ttctgtgatc tcgcggtaca tccaatcagc
    10981 tagctcgatc tcgatgtact ccggccgccc ggtttcgctc tttacgatct tgtagcggct
    11041 aatcaaggct tcaccctcgg ataccgtcac caggcggccg ttcttggcct tcttcgtacg
    11101 ctgcatggca acgtgcgtgg tgtttaaccg aatgcaggtt tctaccaggt cgtctttctg
    11161 ctttccgcca tcggctcgcc ggcagaactt gagtacgtcc gcaacgtgtg gacggaacac
    11221 gcggccgggc ttgtctccct tcccttcccg gtatcggttc atggattcgg ttagatggga
    11281 aaccgccatc agtaccaggt cgtaatccca cacactggcc atgccggccg gccctgcgga
    11341 aacctctacg tgcccgtctg gaagctcgta gcggatcacc tcgccagctc gtcggtcacg
    11401 cttcgacaga cggaaaacgg ccacgtccat gatgctgcga ctatcgcggg tgcccacgtc
    11461 atagagcatc ggaacgaaaa aatctggttg ctcgtcgccc ttgggcggct tcctaatcga
    11521 cggcgcaccg gctgccggcg gttgccggga ttctttgcgg attcgatcag cggccgcttg
    11581 ccacgattca ccggggcgtg cttctgcctc gatgcgttgc cgctgggcgg cctgcgcggc
    11641 cttcaacttc tccaccaggt catcacccag cgccgcgccg atttgtaccg ggccggatgg
    11701 tttgcgaccg ctcacgccga ttcctcgggc ttgggggttc cagtgccatt gcagggccgg
    11761 caggcaaccc agccgcttac gcctggccaa ccgcccgttc ctccacacat ggggcattcc
    11821 acggcgtcgg tgcctggttg ttcttgattt tccatgccgc ctcctttagc cgctaaaatt
    11881 catctactca tttattcatt tgctcattta ctctggtagc tgcgcgatgt attcagatag
    11941 cagctcggta atggtcttgc cttggcgtac cgcgtacatc ttcagcttgg tgtgatcctc
    12001 cgccggcaac tgaaagttga cccgcttcat ggctggcgtg tctgccaggc tggccaacgt
    12061 tgcagccttg ctgctgcgtg cgctcggacg gccggcactt agcgtgtttg tgcttttgct
    12121 cattttctct ttacctcatt aactcaaatg agttttgatt taatttcagc ggccagcgcc
    12181 tggacctcgc gggcagcgtc gccctcgggt tctgattcaa gaacggttgt gccggcggcg
    12241 gcagtgcctg ggtagctcac gcgctgcgtg atacgggact caagaatggg cagctcgtac
    12301 ccggccagcg cctcggcaac ctcaccgccg atgcgcgtgc ctttgatcgc ccgcgacacg
    12361 acaaaggccg cttgtagcct tccatccgtg acctcaatgc gctgcttaac cagctccacc
    12421 aggtcggcgg tggcccatat gtcgtaaggg cttggctgca ccggaatcag cacgaagtcg
    12481 gctgccttga tcgcggacac agccaagtcc gccgcctggg gcgctccgtc gatcactacg
    12541 aagtcgcgcc ggccgatggc cttcacgtcg cggtcaatcg tcgggcggtc gatgccgaca
    12601 acggttagcg gttgatcttc ccgcacggcc gcccaatcgc gggcactgcc ctggggatcg
    12661 gaatcgacta acagaacatc ggccccggcg agttgcaggg cgcgggctag atgggttgcg
    12721 atggtcgtct tgcctgaccc gcctttctgg ttaagtacag cgataacctt catgcgttcc
    12781 ccttgcgtat ttgtttattt actcatcgca tcatatacgc agcgaccgca tgacgcaagc
    12841 tgttttactc aaatacacat caccttttta gacggcggcg ctcggtttct tcagcggcca
    12901 agctggccgg ccaggccgcc agcttggcat cagacaaacc ggccaggatt tcatgcagcc
    12961 gcacggttga gacgtgcgcg ggcggctcga acacgtaccc ggccgcgatc atctccgcct
    13021 cgatctcttc ggtaatgaaa aacggttcgt cctggccgtc ctggtgcggt ttcatgcttg
    13081 ttcctcttgg cgttcattct cggcggccgc cagggcgtcg gcctcggtca atgcgtcctc
    13141 acggaaggca ccgcgccgcc tggcctcggt gggcgtcact tcctcgctgc gctcaagtgc
    13201 gcggtacagg gtcgagcgat gcacgccaag cagtgcagcc gcctctttca cggtgcggcc
    13261 ttcctggtcg atcagctcgc gggcgtgcgc gatctgtgcc ggggtgaggg tagggcgggg
    13321 gccaaacttc acgcctcggg ccttggcggc ctcgcgcccg ctccgggtgc ggtcgatgat
    13381 tagggaacgc tcgaactcgg caatgccggc gaacacggtc aacaccatgc ggccggccgg
    13441 cgtggtggtg tcggcccacg gctctgccag gctacgcagg cccgcgccgg cctcctggat
    13501 gcgctcggca atgtccagta ggtcgcgggt gctgcgggcc aggcggtcta gcctggtcac
    13561 tgtcacaacg tcgccagggc gtaggtggtc aagcatcctg gccagctccg ggcggtcgcg
    13621 cctggtgccg gtgatcttct cggaaaacag cttggtgcag ccggccgcgt gcagttcggc
    13681 ccgttggttg gtcaagtcct ggtcgtcggt gctgacgcgg gcatagccca gcaggccagc
    13741 ggcggcgctc ttgttcatgg cgtaatgtct ccggttctag tcgcaagtat tctactttat
    13801 gcgactaaaa cacgcgacaa gaaaacgcca ggaaaagggc agggcggcag cctgtcgcgt
    13861 aacttaggac ttgtgcgaca tgtcgttttc agaagacggc tgcactgaac gtcagaagcc
    13921 gactgcacta tagcagcgga ggggttggat caaagtactt tgatcccgag gggaaccctg
    13981 tggttggcat gcacatacaa atggacgaac ggataaacct tttcacgccc ttttaaatat
    14041 ccgattattc taataaacgc tcttttctct taggtttacc cgccaatata tcctgtcaaa
    14101 cactgatagt ttaaactgaa ggcgggaaac gacaatctga tccaagctca agctgctcta
    14161 gcattcgcca ttcaggctgc gcaactgttg ggaagggcga tcggtgcggg cctcttcgct
    14221 attacgccag ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg
    14281 gttttcccag tcacgacgtt gtaaaacgac ggccagtgcc aagcttcgac ttgccttccg
    14341 cacaatacat catttcttct tagctttttt tcttcttctt cgttcataca gttttttttt
    14401 gtttatcagc ttacattttc ttgaaccgta gctttcgttt tcttcttttt aactttccat
    14461 tcggagtttt tgtatcttgt ttcatagttt gtcccaggat tagaatgatt aggcatcgaa
    14521 ccttcaagaa tttgattgaa taaaacatct tcattcttaa gatatgaaga taatcttcaa
    14581 aaggcccctg ggaatctgaa agaagagaag caggcccatt tatatgggaa agaacaatag
    14641 tatttcttat ataggcccat ttaagttgaa aacaatcttc aaaagtccca catcgcttag
    14701 ataagaaaac gaagctgagt ttatatacag ctagagtcga agtagtgatt gggagaccaa
    14761 cccagtggac ataagcctgt tcggttcgta agctgtaatg caagtagcgt atgcgctcac
    14821 gcaactggtc cagaaccttg accgaacgca gcggtggtaa cggcgcagtg gcggttttca
    14881 tggcttgtta tgactgtttt tttggggtac agtctatgcc tcgggcatcc aagcagcaag
    14941 cgcgttacgc cgtgggtcga tgtttgatgt tatggagcag caacgatgtt acgcagcagg
    15001 gcagtcgccc taaaacaaag ttaaacatca tgggggaagc ggtgatcgcc gaagtatcga
    15061 ctcaactatc agaggtagtt ggcgtcatcg agcgccatct cgaaccgacg ttgctggccg
    15121 tacatttgta cggctccgca gtggatggcg gcctgaagcc acacagtgat attgatttgc
    15181 tggttacggt gaccgtaagg cttgatgaaa caacgcggcg agctttgatc aacgaccttt
    15241 tggaaacttc ggcttcccct ggagagagcg agattctccg cgctgtagaa gtcaccattg
    15301 ttgtgcacga cgacatcatt ccgtggcgtt atccagctaa gcgcgaactg caatttggag
    15361 aatggcagcg caatgacatt cttgcaggta tcttcgagcc agccacgatc gacattgatc
    15421 tggctatctt gctgacaaaa gcaagagaac atagcgttgc cttggtaggt ccagcggcgg
    15481 aggaactctt tgatccggtt cctgaacagg atctatttga ggcgctaaat gaaaccttaa
    15541 cgctatggaa ctcgccgccc gactgggctg gcgatgagcg aaatgtagtg cttacgttgt
    15601 cccgcatttg gtacagcgca gtaaccggca aaatcgcgcc gaaggatgtc gctgccgact
    15661 gggcaatgga gcgcctgccg gcccagtatc agcccgtcat acttgaagct agacaggctt
    15721 atcttggaca agaagaagat cgcttggcct cgcgcgcaga tcagttggaa gaatttgtcc
    15781 actacgtgaa aggcgagatc accaaggtag tcggcaaata atgtctagct agaaattcgt
    15841 tcaagccgac gccgcttcgc ggcgcggctt aactcaagcg ttagatgcac taagcacata
    15901 attgctcaca gccaaactat caggtcaagt ctgcttttat tatttttaag cgtgcataat
    15961 aagccggtct cggttttaga gctagaaata gcaagttaaa ataaggctag tccgttatca
    16021 acttgaaaaa gtggcaccga gtcggtgctt ttttttgcaa aattttccag atcgatttct
    16081 tcttcctctg ttcttcggcg ttcaatttct ggggttttct cttcgttttc tgtaactgaa
    16141 acctaaaatt tgacctaaaa aaaatctcaa ataatatgat tcagtggttt tgtacttttc
    16201 agttagttga gttttgcagt tccgatgaga taaaccaata ccatggttat actagt
//
