LOCUS       sequence-439115-        9177 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9 and a gRNA targeting C9orf72.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9177)
  AUTHORS   Amick J, Roczniak-Ferguson A, Ferguson SM
  TITLE     C9orf72 binds SMCR8, localizes to lysosomes and regulates mTORC1 
            signaling.
  JOURNAL   Mol Biol Cell. 2016 Aug 24. pii: mbc.E16-01-0003.
  PUBMED    27559131
REFERENCE   2  (bases 1 to 9177)
  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: "Mol Biol 
            Cell. 2016 Aug 24. pii: mbc.E16-01-0003."
COMMENT     Sequence Label: pSpCas9(BB)-2A-Puro-C9orf72
FEATURES             Location/Qualifiers
     source          1..9177
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          76..303
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      315..324
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             324..389
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             396..416
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             441..4541
                     /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             441..4541
                     /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             4542..4589
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4605..4658
                     /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             4659..5258
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(4659..4678)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     complement(5286..5303)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5292..5499
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5508..5648
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5508..5637
                     /label=AAV2 ITR
     rep_origin      5723..6178
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5810..5829)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6020..6041
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6195..6214)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6314..6336
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6374..6392)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6460..6564
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6565..6633,6634..7425)
                     /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     complement(6783..6802)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7596..8184
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8085..8104
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        8246..8494
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     8246..8266
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     8417..8436
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        8516..8591
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        8688..8973
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        join(8975..9177,1..75)
                     /label=chicken beta-actin promoter
ORIGIN
        1 gcgctccgaa agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga
       61 agcgcgcggc gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc
      121 gcctcgcgcc gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg
      181 gacggccctt ctcctccggg ctgtaattag ctgagcaaga ggtaagggtt taagggatgg
      241 ttggttggtg gggtattaat gtttaattac ctggagcacc tgcctgaaat cacttttttt
      301 caggttggac cggtgccacc atggactata aggaccacga cggagactac aaggatcatg
      361 atattgatta caaagacgat gacgataaga tggccccaaa gaagaagcgg aaggtcggta
      421 tccacggagt cccagcagcc gacaagaagt acagcatcgg cctggacatc ggcaccaact
      481 ctgtgggctg ggccgtgatc accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc
      541 tgggcaacac cgaccggcac agcatcaaga agaacctgat cggagccctg ctgttcgaca
      601 gcggcgaaac agccgaggcc acccggctga agagaaccgc cagaagaaga tacaccagac
      661 ggaagaaccg gatctgctat ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg
      721 acagcttctt ccacagactg gaagagtcct tcctggtgga agaggataag aagcacgagc
      781 ggcaccccat cttcggcaac atcgtggacg aggtggccta ccacgagaag taccccacca
      841 tctaccacct gagaaagaaa ctggtggaca gcaccgacaa ggccgacctg cggctgatct
      901 atctggccct ggcccacatg atcaagttcc ggggccactt cctgatcgag ggcgacctga
      961 accccgacaa cagcgacgtg gacaagctgt tcatccagct ggtgcagacc tacaaccagc
     1021 tgttcgagga aaaccccatc aacgccagcg gcgtggacgc caaggccatc ctgtctgcca
     1081 gactgagcaa gagcagacgg ctggaaaatc tgatcgccca gctgcccggc gagaagaaga
     1141 atggcctgtt cggaaacctg attgccctga gcctgggcct gacccccaac ttcaagagca
     1201 acttcgacct ggccgaggat gccaaactgc agctgagcaa ggacacctac gacgacgacc
     1261 tggacaacct gctggcccag atcggcgacc agtacgccga cctgtttctg gccgccaaga
     1321 acctgtccga cgccatcctg ctgagcgaca tcctgagagt gaacaccgag atcaccaagg
     1381 cccccctgag cgcctctatg atcaagagat acgacgagca ccaccaggac ctgaccctgc
     1441 tgaaagctct cgtgcggcag cagctgcctg agaagtacaa agagattttc ttcgaccaga
     1501 gcaagaacgg ctacgccggc tacattgacg gcggagccag ccaggaagag ttctacaagt
     1561 tcatcaagcc catcctggaa aagatggacg gcaccgagga actgctcgtg aagctgaaca
     1621 gagaggacct gctgcggaag cagcggacct tcgacaacgg cagcatcccc caccagatcc
     1681 acctgggaga gctgcacgcc attctgcggc ggcaggaaga tttttaccca ttcctgaagg
     1741 acaaccggga aaagatcgag aagatcctga ccttccgcat cccctactac gtgggccctc
     1801 tggccagggg aaacagcaga ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc
     1861 cctggaactt cgaggaagtg gtggacaagg gcgcttccgc ccagagcttc atcgagcgga
     1921 tgaccaactt cgataagaac ctgcccaacg agaaggtgct gcccaagcac agcctgctgt
     1981 acgagtactt caccgtgtat aacgagctga ccaaagtgaa atacgtgacc gagggaatga
     2041 gaaagcccgc cttcctgagc ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga
     2101 ccaaccggaa agtgaccgtg aagcagctga aagaggacta cttcaagaaa atcgagtgct
     2161 tcgactccgt ggaaatctcc ggcgtggaag atcggttcaa cgcctccctg ggcacatacc
     2221 acgatctgct gaaaattatc aaggacaagg acttcctgga caatgaggaa aacgaggaca
     2281 ttctggaaga tatcgtgctg accctgacac tgtttgagga cagagagatg atcgaggaac
     2341 ggctgaaaac ctatgcccac ctgttcgacg acaaagtgat gaagcagctg aagcggcgga
     2401 gatacaccgg ctggggcagg ctgagccgga agctgatcaa cggcatccgg gacaagcagt
     2461 ccggcaagac aatcctggat ttcctgaagt ccgacggctt cgccaacaga aacttcatgc
     2521 agctgatcca cgacgacagc ctgaccttta aagaggacat ccagaaagcc caggtgtccg
     2581 gccagggcga tagcctgcac gagcacattg ccaatctggc cggcagcccc gccattaaga
     2641 agggcatcct gcagacagtg aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca
     2701 agcccgagaa catcgtgatc gaaatggcca gagagaacca gaccacccag aagggacaga
     2761 agaacagccg cgagagaatg aagcggatcg aagagggcat caaagagctg ggcagccaga
     2821 tcctgaaaga acaccccgtg gaaaacaccc agctgcagaa cgagaagctg tacctgtact
     2881 acctgcagaa tgggcgggat atgtacgtgg accaggaact ggacatcaac cggctgtccg
     2941 actacgatgt ggaccatatc gtgcctcaga gctttctgaa ggacgactcc atcgacaaca
     3001 aggtgctgac cagaagcgac aagaaccggg gcaagagcga caacgtgccc tccgaagagg
     3061 tcgtgaagaa gatgaagaac tactggcggc agctgctgaa cgccaagctg attacccaga
     3121 gaaagttcga caatctgacc aaggccgaga gaggcggcct gagcgaactg gataaggccg
     3181 gcttcatcaa gagacagctg gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc
     3241 tggactcccg gatgaacact aagtacgacg agaatgacaa gctgatccgg gaagtgaaag
     3301 tgatcaccct gaagtccaag ctggtgtccg atttccggaa ggatttccag ttttacaaag
     3361 tgcgcgagat caacaactac caccacgccc acgacgccta cctgaacgcc gtcgtgggaa
     3421 ccgccctgat caaaaagtac cctaagctgg aaagcgagtt cgtgtacggc gactacaagg
     3481 tgtacgacgt gcggaagatg atcgccaaga gcgagcagga aatcggcaag gctaccgcca
     3541 agtacttctt ctacagcaac atcatgaact ttttcaagac cgagattacc ctggccaacg
     3601 gcgagatccg gaagcggcct ctgatcgaga caaacggcga aaccggggag atcgtgtggg
     3661 ataagggccg ggattttgcc accgtgcgga aagtgctgag catgccccaa gtgaatatcg
     3721 tgaaaaagac cgaggtgcag acaggcggct tcagcaaaga gtctatcctg cccaagagga
     3781 acagcgataa gctgatcgcc agaaagaagg actgggaccc taagaagtac ggcggcttcg
     3841 acagccccac cgtggcctat tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca
     3901 agaaactgaa gagtgtgaaa gagctgctgg ggatcaccat catggaaaga agcagcttcg
     3961 agaagaatcc catcgacttt ctggaagcca agggctacaa agaagtgaaa aaggacctga
     4021 tcatcaagct gcctaagtac tccctgttcg agctggaaaa cggccggaag agaatgctgg
     4081 cctctgccgg cgaactgcag aagggaaacg aactggccct gccctccaaa tatgtgaact
     4141 tcctgtacct ggccagccac tatgagaagc tgaagggctc ccccgaggat aatgagcaga
     4201 aacagctgtt tgtggaacag cacaagcact acctggacga gatcatcgag cagatcagcg
     4261 agttctccaa gagagtgatc ctggccgacg ctaatctgga caaagtgctg tccgcctaca
     4321 acaagcaccg ggataagccc atcagagagc aggccgagaa tatcatccac ctgtttaccc
     4381 tgaccaatct gggagcccct gccgccttca agtactttga caccaccatc gaccggaaga
     4441 ggtacaccag caccaaagag gtgctggacg ccaccctgat ccaccagagc atcaccggcc
     4501 tgtacgagac acggatcgac ctgtctcagc tgggaggcga caaaaggccg gcggccacga
     4561 aaaaggccgg ccaggcaaaa aagaaaaagg aattcggcag tggagagggc agaggaagtc
     4621 tgctaacatg cggtgacgtc gaggagaatc ctggcccaat gaccgagtac aagcccacgg
     4681 tgcgcctcgc cacccgcgac gacgtcccca gggccgtacg caccctcgcc gccgcgttcg
     4741 ccgactaccc cgccacgcgc cacaccgtcg atccggaccg ccacatcgag cgggtcaccg
     4801 agctgcaaga actcttcctc acgcgcgtcg ggctcgacat cggcaaggtg tgggtcgcgg
     4861 acgacggcgc cgcggtggcg gtctggacca cgccggagag cgtcgaagcg ggggcggtgt
     4921 tcgccgagat cggcccgcgc atggccgagt tgagcggttc ccggctggcc gcgcagcaac
     4981 agatggaagg cctcctggcg ccgcaccggc ccaaggagcc cgcgtggttc ctggccaccg
     5041 tcggcgtctc gcccgaccac cagggcaagg gtctgggcag cgccgtcgtg ctccccggag
     5101 tggaggcggc cgagcgcgcc ggggtgcccg ccttcctgga gacctccgcg ccccacaacc
     5161 tccccttcta cgagcggctc ggcttcaccg tcaccgccga cgtcgaggtg cccgaaggac
     5221 cgcgcacctg gtgcatgacc cgcaagcccg gtgcctgaga attctaacta gagctcgctg
     5281 atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc
     5341 ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc
     5401 atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa
     5461 gggggaggat tgggaagaga atagcaggca tgctggggag cggccgcagg aacccctagt
     5521 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     5581 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     5641 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     5701 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     5761 gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct
     5821 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     5881 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     5941 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     6001 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc
     6061 tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat
     6121 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     6181 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     6241 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     6301 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     6361 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     6421 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     6481 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     6541 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     6601 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     6661 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     6721 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     6781 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     6841 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     6901 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     6961 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     7021 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     7081 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     7141 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     7201 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     7261 tctggagccg gtgagcgtgg aagccgcggt atcattgcag cactggggcc agatggtaag
     7321 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     7381 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     7441 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     7501 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     7561 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     7621 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     7681 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     7741 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     7801 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     7861 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     7921 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     7981 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     8041 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     8101 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     8161 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     8221 ttttgctggc cttttgctca catgtgaggg cctatttccc atgattcctt catatttgca
     8281 tatacgatac aaggctgtta gagagataat tggaattaat ttgactgtaa acacaaagat
     8341 attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa
     8401 attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt
     8461 ggctttatat atcttgtgga aaggacgaaa caccggctta ctgggacaat attctgtttt
     8521 agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa aaagtggcac
     8581 cgagtcggtg cttttttgtt ttagagctag aaatagcaag ttaaaataag gctagtccgt
     8641 ttttagcgcg tgcgccaatt ctgcagacaa atggctctag aggtacccgt tacataactt
     8701 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaatagta
     8761 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     8821 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
     8881 aaatggcccg cctggcattg tgcccagtac atgaccttat gggactttcc tacttggcag
     8941 tacatctacg tattagtcat cgctattacc atggtcgagg tgagccccac gttctgcttc
     9001 actctcccca tctccccccc ctccccaccc ccaattttgt atttatttat tttttaatta
     9061 ttttgtgcag cgatgggggc gggggggggg ggggggcgcg cgccaggcgg ggcggggcgg
     9121 ggcgaggggc ggggcggggc gaggcggaga ggtgcggcgg cagccaatca gagcggc
//
