LOCUS       sequence-378800-        9117 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For CRISPR knockout of integrin alpha5.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9117)
  AUTHORS   Lin FY, Li J, Xie Y, Zhu J, Huong Nguyen TT, Zhang Y, Zhu J, 
            Springer TA
  TITLE     A general chemical principle for creating closure-stabilizing 
            integrin inhibitors.
  JOURNAL   Cell. 2022 Sep 15;185(19):3533-3550.e27. doi: 
            10.1016/j.cell.2022.08.008.
  PUBMED    36113427
REFERENCE   2  (bases 1 to 9117)
  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"; doi: 
            "10.1016/j.cell.2022.08"; journalName: "Cell"; date: "2022-09-15- 
            15"; volume: "185"; issue: "19"; pages: "3533-3550.e27"
COMMENT     Sequence Label: pSpCas9n(BB)-2A-puro-a5 guide A
FEATURES             Location/Qualifiers
     source          1..9117
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          103..330
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      342..351
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             351..416
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             423..443
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(468..491,492..494,495..4568)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             468..4568
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             4569..4616
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             4632..4685
                     /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             4686..5285
                     /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(4686..4705)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     5182..5202
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     primer_bind     complement(5313..5330)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5319..5526
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5535..5675
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5535..5664
                     /label=AAV2 ITR
     rep_origin      5750..6205
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5837..5856)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6047..6068
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6222..6241)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6341..6363
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6401..6419)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6487..6591
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6592..6660,6661..7452)
                     /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(6810..6829)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7623..8211
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8112..8131
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        8273..8521
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     8273..8293
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     8444..8463
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        8543..8618
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        8715..9000
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
ORIGIN
        1 ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg
       61 cggcggcggc ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc
      121 cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc
      181 gcgttactcc cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagctg
      241 agcaagaggt aagggtttaa gggatggttg gttggtgggg tattaatgtt taattacctg
      301 gagcacctgc ctgaaatcac tttttttcag gttggaccgg tgccaccatg gactataagg
      361 accacgacgg agactacaag gatcatgata ttgattacaa agacgatgac gataagatgg
      421 ccccaaagaa gaagcggaag gtcggtatcc acggagtccc agcagccgac aagaagtaca
      481 gcatcggcct ggccatcggc accaactctg tgggctgggc cgtgatcacc gacgagtaca
      541 aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc atcaagaaga
      601 acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga
      661 gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg caagagatct
      721 tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa gagtccttcc
      781 tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc gtggacgagg
      841 tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg gtggacagca
      901 ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc aagttccggg
      961 gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac aagctgttca
     1021 tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg
     1081 tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg gaaaatctga
     1141 tcgcccagct gcccggcgag aagaagaatg gcctgttcgg caacctgatt gccctgagcc
     1201 tgggcctgac ccccaacttc aagagcaact tcgacctggc cgaggatgcc aaactgcagc
     1261 tgagcaagga cacctacgac gacgacctgg acaacctgct ggcccagatc ggcgaccagt
     1321 acgccgacct gtttctggcc gccaagaacc tgtccgacgc catcctgctg agcgacatcc
     1381 tgagagtgaa caccgagatc accaaggccc ccctgagcgc ctctatgatc aagagatacg
     1441 acgagcacca ccaggacctg accctgctga aagctctcgt gcggcagcag ctgcctgaga
     1501 agtacaaaga gattttcttc gaccagagca agaacggcta cgccggctac attgacggcg
     1561 gagccagcca ggaagagttc tacaagttca tcaagcccat cctggaaaag atggacggca
     1621 ccgaggaact gctcgtgaag ctgaacagag aggacctgct gcggaagcag cggaccttcg
     1681 acaacggcag catcccccac cagatccacc tgggagagct gcacgccatt ctgcggcggc
     1741 aggaagattt ttacccattc ctgaaggaca accgggaaaa gatcgagaag atcctgacct
     1801 tccgcatccc ctactacgtg ggccctctgg ccaggggaaa cagcagattc gcctggatga
     1861 ccagaaagag cgaggaaacc atcaccccct ggaacttcga ggaagtggtg gacaagggcg
     1921 cttccgccca gagcttcatc gagcggatga ccaacttcga taagaacctg cccaacgaga
     1981 aggtgctgcc caagcacagc ctgctgtacg agtacttcac cgtgtataac gagctgacca
     2041 aagtgaaata cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa
     2101 aggccatcgt ggacctgctg ttcaagacca accggaaagt gaccgtgaag cagctgaaag
     2161 aggactactt caagaaaatc gagtgcttcg actccgtgga aatctccggc gtggaagatc
     2221 ggttcaacgc ctccctgggc acataccacg atctgctgaa aattatcaag gacaaggact
     2281 tcctggacaa tgaggaaaac gaggacattc tggaagatat cgtgctgacc ctgacactgt
     2341 ttgaggacag agagatgatc gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca
     2401 aagtgatgaa gcagctgaag cggcggagat acaccggctg gggcaggctg agccggaagc
     2461 tgatcaacgg catccgggac aagcagtccg gcaagacaat cctggatttc ctgaagtccg
     2521 acggcttcgc caacagaaac ttcatgcagc tgatccacga cgacagcctg acctttaaag
     2581 aggacatcca gaaagcccag gtgtccggcc agggcgatag cctgcacgag cacattgcca
     2641 atctggccgg cagccccgcc attaagaagg gcatcctgca gacagtgaag gtggtggacg
     2701 agctcgtgaa agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa atggccagag
     2761 agaaccagac cacccagaag ggacagaaga acagccgcga gagaatgaag cggatcgaag
     2821 agggcatcaa agagctgggc agccagatcc tgaaagaaca ccccgtggaa aacacccagc
     2881 tgcagaacga gaagctgtac ctgtactacc tgcagaatgg gcgggatatg tacgtggacc
     2941 aggaactgga catcaaccgg ctgtccgact acgatgtgga ccatatcgtg cctcagagct
     3001 ttctgaagga cgactccatc gacaacaagg tgctgaccag aagcgacaag aaccggggca
     3061 agagcgacaa cgtgccctcc gaagaggtcg tgaagaagat gaagaactac tggcggcagc
     3121 tgctgaacgc caagctgatt acccagagaa agttcgacaa tctgaccaag gccgagagag
     3181 gcggcctgag cgaactggat aaggccggct tcatcaagag acagctggtg gaaacccggc
     3241 agatcacaaa gcacgtggca cagatcctgg actcccggat gaacactaag tacgacgaga
     3301 atgacaagct gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt
     3361 tccggaagga tttccagttt tacaaagtgc gcgagatcaa caactaccac cacgcccacg
     3421 acgcctacct gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa
     3481 gcgagttcgt gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg
     3541 agcaggaaat cggcaaggct accgccaagt acttcttcta cagcaacatc atgaactttt
     3601 tcaagaccga gattaccctg gccaacggcg agatccggaa gcggcctctg atcgagacaa
     3661 acggcgaaac cggggagatc gtgtgggata agggccggga ttttgccacc gtgcggaaag
     3721 tgctgagcat gccccaagtg aatatcgtga aaaagaccga ggtgcagaca ggcggcttca
     3781 gcaaagagtc tatcctgccc aagaggaaca gcgataagct gatcgccaga aagaaggact
     3841 gggaccctaa gaagtacggc ggcttcgaca gccccaccgt ggcctattct gtgctggtgg
     3901 tggccaaagt ggaaaagggc aagtccaaga aactgaagag tgtgaaagag ctgctgggga
     3961 tcaccatcat ggaaagaagc agcttcgaga agaatcccat cgactttctg gaagccaagg
     4021 gctacaaaga agtgaaaaag gacctgatca tcaagctgcc taagtactcc ctgttcgagc
     4081 tggaaaacgg ccggaagaga atgctggcct ctgccggcga actgcagaag ggaaacgaac
     4141 tggccctgcc ctccaaatat gtgaacttcc tgtacctggc cagccactat gagaagctga
     4201 agggctcccc cgaggataat gagcagaaac agctgtttgt ggaacagcac aagcactacc
     4261 tggacgagat catcgagcag atcagcgagt tctccaagag agtgatcctg gccgacgcta
     4321 atctggacaa agtgctgtcc gcctacaaca agcaccggga taagcccatc agagagcagg
     4381 ccgagaatat catccacctg tttaccctga ccaatctggg agcccctgcc gccttcaagt
     4441 actttgacac caccatcgac cggaagaggt acaccagcac caaagaggtg ctggacgcca
     4501 ccctgatcca ccagagcatc accggcctgt acgagacacg gatcgacctg tctcagctgg
     4561 gaggcgacaa aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaaggaat
     4621 tcggcagtgg agagggcaga ggaagtctgc taacatgcgg tgacgtcgag gagaatcctg
     4681 gcccaatgac cgagtacaag cccacggtgc gcctcgccac ccgcgacgac gtccccaggg
     4741 ccgtacgcac cctcgccgcc gcgttcgccg actaccccgc cacgcgccac accgtcgatc
     4801 cggaccgcca catcgagcgg gtcaccgagc tgcaagaact cttcctcacg cgcgtcgggc
     4861 tcgacatcgg caaggtgtgg gtcgcggacg acggcgccgc ggtggcggtc tggaccacgc
     4921 cggagagcgt cgaagcgggg gcggtgttcg ccgagatcgg cccgcgcatg gccgagttga
     4981 gcggttcccg gctggccgcg cagcaacaga tggaaggcct cctggcgccg caccggccca
     5041 aggagcccgc gtggttcctg gccaccgtcg gagtctcgcc cgaccaccag ggcaagggtc
     5101 tgggcagcgc cgtcgtgctc cccggagtgg aggcggccga gcgcgccggg gtgcccgcct
     5161 tcctggagac ctccgcgccc cgcaacctcc ccttctacga gcggctcggc ttcaccgtca
     5221 ccgccgacgt cgaggtgccc gaaggaccgc gcacctggtg catgacccgc aagcccggtg
     5281 cctgagaatt ctaactagag ctcgctgatc agcctcgact gtgccttcta gttgccagcc
     5341 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     5401 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     5461 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagagaata gcaggcatgc
     5521 tggggagcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc tgcgcgctcg
     5581 ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc
     5641 ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt attttctcct
     5701 tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta cgcgccctgt
     5761 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     5821 agcgccttag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     5881 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
     5941 cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc atcgccctga
     6001 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
     6061 caaactggaa caacactcaa ctctatctcg ggctattctt ttgatttata agggattttg
     6121 ccgatttcgg tctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
     6181 aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg ctctgatgcc
     6241 gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg acgggcttgt
     6301 ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag
     6361 aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt
     6421 ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac ttttcgggga
     6481 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     6541 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     6601 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     6661 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     6721 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     6781 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     6841 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     6901 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     6961 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     7021 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     7081 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     7141 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     7201 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     7261 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtggaag ccgcggtatc
     7321 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     7381 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     7441 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     7501 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     7561 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     7621 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
     7681 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
     7741 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
     7801 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
     7861 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
     7921 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
     7981 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
     8041 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
     8101 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
     8161 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
     8221 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gtgagggcct
     8281 atttcccatg attccttcat atttgcatat acgatacaag gctgttagag agataattgg
     8341 aattaatttg actgtaaaca caaagatatt agtacaaaat acgtgacgta gaaagtaata
     8401 atttcttggg tagtttgcag ttttaaaatt atgttttaaa atggactatc atatgcttac
     8461 cgtaacttga aagtatttcg atttcttggc tttatatatc ttgtggaaag gacgaaacac
     8521 cggtacagtg gagcgcatgc ctgttttaga gctagaaata gcaagttaaa ataaggctag
     8581 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt ttttgtttta gagctagaaa
     8641 tagcaagtta aaataaggct agtccgtttt tagcgcgtgc gccaattctg cagacaaatg
     8701 gctctagagg tacccgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
     8761 gacccccgcc cattgacgtc aatagtaacg ccaataggga ctttccattg acgtcaatgg
     8821 gtggagtatt tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt
     8881 acgcccccta ttgacgtcaa tgacggtaaa tggcccgcct ggcattgtgc ccagtacatg
     8941 accttatggg actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg
     9001 gtcgaggtga gccccacgtt ctgcttcact ctccccatct cccccccctc cccaccccca
     9061 attttgtatt tatttatttt ttaattattt tgtgcagcga tgggggcggg ggggggg
//
