LOCUS       sequence-439116-        9178 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9 and a gRNA targeting SMCR8.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9178)
  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 9178)
  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-SMCR8
FEATURES             Location/Qualifiers
     source          1..9178
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3699..3746
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             3762..3815
                     /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             3816..4415
                     /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(3816..3835)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     complement(4443..4460)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4449..4656
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4665..4805
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4665..4794
                     /label=AAV2 ITR
     rep_origin      4880..5335
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4967..4986)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5177..5198
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5352..5371)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5471..5493
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5531..5549)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5617..5721
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5722..5790,5791..6582)
                     /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(5940..5959)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6753..7341
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7242..7261
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        7403..7651
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7403..7423
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7574..7593
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        7673..7748
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        7845..8130
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        8132..8410
                     /label=chicken beta-actin promoter
     intron          8411..8638
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      8650..8659
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             8659..8724
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8731..8751
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(8776..9178,1..3698)
                     /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             join(8776..9178,1..3698)
                     /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"
ORIGIN
        1 accacctgag aaagaaactg gtggacagca ccgacaaggc cgacctgcgg ctgatctatc
       61 tggccctggc ccacatgatc aagttccggg gccacttcct gatcgagggc gacctgaacc
      121 ccgacaacag cgacgtggac aagctgttca tccagctggt gcagacctac aaccagctgt
      181 tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa ggccatcctg tctgccagac
      241 tgagcaagag cagacggctg gaaaatctga tcgcccagct gcccggcgag aagaagaatg
      301 gcctgttcgg aaacctgatt gccctgagcc tgggcctgac ccccaacttc aagagcaact
      361 tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac gacgacctgg
      421 acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc gccaagaacc
      481 tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc accaaggccc
      541 ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg accctgctga
      601 aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc gaccagagca
      661 agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc tacaagttca
      721 tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag ctgaacagag
      781 aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac cagatccacc
      841 tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc ctgaaggaca
      901 accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg ggccctctgg
      961 ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc atcaccccct
     1021 ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc gagcggatga
     1081 ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc ctgctgtacg
     1141 agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag ggaatgagaa
     1201 agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg ttcaagacca
     1261 accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc gagtgcttcg
     1321 actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc acataccacg
     1381 atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac gaggacattc
     1441 tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc gaggaacggc
     1501 tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag cggcggagat
     1561 acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac aagcagtccg
     1621 gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac ttcatgcagc
     1681 tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag gtgtccggcc
     1741 agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc attaagaagg
     1801 gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc cggcacaagc
     1861 ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag ggacagaaga
     1921 acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc agccagatcc
     1981 tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac ctgtactacc
     2041 tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg ctgtccgact
     2101 acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc gacaacaagg
     2161 tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc gaagaggtcg
     2221 tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt acccagagaa
     2281 agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat aaggccggct
     2341 tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca cagatcctgg
     2401 actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa gtgaaagtga
     2461 tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt tacaaagtgc
     2521 gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc gtgggaaccg
     2581 ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac tacaaggtgt
     2641 acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct accgccaagt
     2701 acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg gccaacggcg
     2761 agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc gtgtgggata
     2821 agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg aatatcgtga
     2881 aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc aagaggaaca
     2941 gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc ggcttcgaca
     3001 gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc aagtccaaga
     3061 aactgaagag tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc agcttcgaga
     3121 agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag gacctgatca
     3181 tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga atgctggcct
     3241 ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat gtgaacttcc
     3301 tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat gagcagaaac
     3361 agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag atcagcgagt
     3421 tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc gcctacaaca
     3481 agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg tttaccctga
     3541 ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac cggaagaggt
     3601 acaccagcac caaagaggtg ctggacgcca ccctgatcca ccagagcatc accggcctgt
     3661 acgagacacg gatcgacctg tctcagctgg gaggcgacaa aaggccggcg gccacgaaaa
     3721 aggccggcca ggcaaaaaag aaaaaggaat tcggcagtgg agagggcaga ggaagtctgc
     3781 taacatgcgg tgacgtcgag gagaatcctg gcccaatgac cgagtacaag cccacggtgc
     3841 gcctcgccac ccgcgacgac gtccccaggg ccgtacgcac cctcgccgcc gcgttcgccg
     3901 actaccccgc cacgcgccac accgtcgatc cggaccgcca catcgagcgg gtcaccgagc
     3961 tgcaagaact cttcctcacg cgcgtcgggc tcgacatcgg caaggtgtgg gtcgcggacg
     4021 acggcgccgc ggtggcggtc tggaccacgc cggagagcgt cgaagcgggg gcggtgttcg
     4081 ccgagatcgg cccgcgcatg gccgagttga gcggttcccg gctggccgcg cagcaacaga
     4141 tggaaggcct cctggcgccg caccggccca aggagcccgc gtggttcctg gccaccgtcg
     4201 gcgtctcgcc cgaccaccag ggcaagggtc tgggcagcgc cgtcgtgctc cccggagtgg
     4261 aggcggccga gcgcgccggg gtgcccgcct tcctggagac ctccgcgccc cacaacctcc
     4321 ccttctacga gcggctcggc ttcaccgtca ccgccgacgt cgaggtgccc gaaggaccgc
     4381 gcacctggtg catgacccgc aagcccggtg cctgagaatt ctaactagag ctcgctgatc
     4441 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     4501 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     4561 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     4621 ggaggattgg gaagagaata gcaggcatgc tggggagcgg ccgcaggaac ccctagtgat
     4681 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     4741 cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct
     4801 gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     4861 acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     4921 gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc tttcgctttc
     4981 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
     5041 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgatttgggt
     5101 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
     5161 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ctctatctcg
     5221 ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt aaaaaatgag
     5281 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac aattttatgg
     5341 tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg acacccgcca
     5401 acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta cagacaagct
     5461 gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg
     5521 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
     5581 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     5641 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     5701 taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     5761 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     5821 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     5881 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     5941 ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg tcgccgcata
     6001 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     6061 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     6121 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     6181 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     6241 gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa actattaact
     6301 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     6361 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     6421 ggagccggtg agcgtggaag ccgcggtatc attgcagcac tggggccaga tggtaagccc
     6481 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     6541 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga ccaagtttac
     6601 tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat ctaggtgaag
     6661 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     6721 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     6781 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     6841 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc aaatactgtt
     6901 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     6961 ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc gtgtcttacc
     7021 gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg aacggggggt
     7081 tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata cctacagcgt
     7141 gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta tccggtaagc
     7201 ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc ctggtatctt
     7261 tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg atgctcgtca
     7321 ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt cctggccttt
     7381 tgctggcctt ttgctcacat gtgagggcct atttcccatg attccttcat atttgcatat
     7441 acgatacaag gctgttagag agataattgg aattaatttg actgtaaaca caaagatatt
     7501 agtacaaaat acgtgacgta gaaagtaata atttcttggg tagtttgcag ttttaaaatt
     7561 atgttttaaa atggactatc atatgcttac cgtaacttga aagtatttcg atttcttggc
     7621 tttatatatc ttgtggaaag gacgaaacac cggatcagcg cccctgacgt aggttttaga
     7681 gctagaaata gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga
     7741 gtcggtgctt ttttgtttta gagctagaaa tagcaagtta aaataaggct agtccgtttt
     7801 tagcgcgtgc gccaattctg cagacaaatg gctctagagg tacccgttac ataacttacg
     7861 gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc aatagtaacg
     7921 ccaataggga ctttccattg acgtcaatgg gtggagtatt tacggtaaac tgcccacttg
     7981 gcagtacatc aagtgtatca tatgccaagt acgcccccta ttgacgtcaa tgacggtaaa
     8041 tggcccgcct ggcattgtgc ccagtacatg accttatggg actttcctac ttggcagtac
     8101 atctacgtat tagtcatcgc tattaccatg gtcgaggtga gccccacgtt ctgcttcact
     8161 ctccccatct cccccccctc cccaccccca attttgtatt tatttatttt ttaattattt
     8221 tgtgcagcga tgggggcggg gggggggggg ggggcgcgcg ccaggcgggg cggggcgggg
     8281 cgaggggcgg ggcggggcga ggcggagagg tgcggcggca gccaatcaga gcggcgcgct
     8341 ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg ccctataaaa agcgaagcgc
     8401 gcggcgggcg ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc
     8461 gcgccgcccg ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg
     8521 cccttctcct ccgggctgta attagctgag caagaggtaa gggtttaagg gatggttggt
     8581 tggtggggta ttaatgttta attacctgga gcacctgcct gaaatcactt tttttcaggt
     8641 tggaccggtg ccaccatgga ctataaggac cacgacggag actacaagga tcatgatatt
     8701 gattacaaag acgatgacga taagatggcc ccaaagaaga agcggaaggt cggtatccac
     8761 ggagtcccag cagccgacaa gaagtacagc atcggcctgg acatcggcac caactctgtg
     8821 ggctgggccg tgatcaccga cgagtacaag gtgcccagca agaaattcaa ggtgctgggc
     8881 aacaccgacc ggcacagcat caagaagaac ctgatcggag ccctgctgtt cgacagcggc
     8941 gaaacagccg aggccacccg gctgaagaga accgccagaa gaagatacac cagacggaag
     9001 aaccggatct gctatctgca agagatcttc agcaacgaga tggccaaggt ggacgacagc
     9061 ttcttccaca gactggaaga gtccttcctg gtggaagagg ataagaagca cgagcggcac
     9121 cccatcttcg gcaacatcgt ggacgaggtg gcctaccacg agaagtaccc caccatct
//
