LOCUS       sequence-245464-       10467 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Expresses SpyCas9 fused to attenuated AcrIIA4 (insertion 5).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10467)
  AUTHORS   Aschenbrenner S, Kallenberger SM, Hoffmann MD, Huck A, Eils R, 
            Niopek D
  TITLE     Coupling Cas9 to artificial inhibitory domains enhances CRISPR-Cas9 
            target specificity.
  JOURNAL   Sci Adv. 2020 Feb 5;6(6):eaay0187. doi: 10.1126/sciadv.aay0187. 
            eCollection 2020 Feb.
  PUBMED    32076642
REFERENCE   2  (bases 1 to 10467)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv."; 
            date: "2020-02-5"; volume: "6(6):eaay0187. doi"; pages: " 
            10.1126/sciadv.aay0187. eCollection 2020 Feb"
COMMENT     Sequence Label: Cas-Acr (insertion 5)
FEATURES             Location/Qualifiers
     source          1..10467
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      931..940
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             940..1005
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1012..1032
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1057..5157
                     /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             1057..5157
                     /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             5158..5205
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(6061..6078)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6067..6291
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      6337..6765
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6424..6443)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6634..6655
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6774..6794)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        6779..7108
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6959..7094
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7021..7040
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             7175..7969
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(7229..7248)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7839..7858
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    8143..8264
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8180..8199)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8234..8253
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8313..8329)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8313..8329)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8326..8348)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8337..8353
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(8361..8367,8368..8385,8386..8391))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8406..8427
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8544..8561)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8715..9300)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8795..8814)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9471..10262,10263..10331))
                     /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     10094..10113
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10332..10436)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaacgg gccctctaga ctcgagcggt gccaccatgg actataagga ccacgacgga
      961 gactacaagg atcatgatat tgattacaaa gacgatgacg ataagatggc cccaaagaag
     1021 aagcggaagg tcggtatcca cggagtccca gcagccgaca agaagtacag catcggcctg
     1081 gacatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc
     1141 aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcgga
     1201 gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag aaccgccaga
     1261 agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag
     1321 atggccaagg tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag
     1381 gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac
     1441 gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc
     1501 gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg
     1561 atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg
     1621 cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag
     1681 gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg
     1741 cccggcgaga agaagaatgg cctgttcgga aacctgattg ccctgagcct gggcctgacc
     1801 cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac
     1861 acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg
     1921 tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac
     1981 accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac
     2041 caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag
     2101 attttcttcg accagagcaa gaacggctac gccggctaca ttgacggcgg agccagccag
     2161 gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg
     2221 ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc
     2281 atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt
     2341 tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc
     2401 tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc
     2461 gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc ttccgcccag
     2521 agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc
     2581 aagcacagcc tgctgtacga gtacttcacc gtgtataacg agctgaccaa agtgaaatac
     2641 gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg
     2701 gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc
     2761 aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc
     2821 tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat
     2881 gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga
     2941 gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag
     3001 cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc
     3061 atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc
     3121 aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag
     3181 aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc
     3241 agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa
     3301 gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc
     3361 acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa
     3421 gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag
     3481 aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac
     3541 atcaaccggc tgtccgacta cgatgtggac catatcgtgc ctcagagctt tctgaaggac
     3601 gactccatcg acaacaaggt gctgaccaga agcgacaaga accggggcaa gagcgacaac
     3661 gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct gctgaacgcc
     3721 aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc
     3781 gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag
     3841 cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa tgacaagctg
     3901 atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat
     3961 ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg
     4021 aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg
     4081 tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc
     4141 ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag
     4201 attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaacc
     4261 ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt gctgagcatg
     4321 ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct
     4381 atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg ggaccctaag
     4441 aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg
     4501 gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg
     4561 gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa
     4621 gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc
     4681 cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc
     4741 tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc
     4801 gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct ggacgagatc
     4861 atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa
     4921 gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc cgagaatatc
     4981 atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc
     5041 accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac
     5101 cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacaaa
     5161 aggccggcgg ccacgaaaaa ggccggccag gcaaaaaaga aaaaggaatt cgggggatca
     5221 gggggagggt ctggtggtgg atctggtgga ggatctggtg ggggttcagg gggcggttcc
     5281 ggaggaggct caggaggcgg ctccggcggt ggctcaggag gtggatccgg tatgaacatt
     5341 aacgacctga ttcgggaaat aaagaacaag gactacacgg tcaagctgtc aggcaccgac
     5401 tccaactcta taacacagct cattataagg gttaataatg atggcaacga atatgtcatc
     5461 tctgaatccg agaatgaatc aatcgtggag aaattcatca gtgcatttaa aaacggctgg
     5521 cttgctgcag cgttggagcg gattgagaag aatttcgtca taactgatcc caggttgcct
     5581 gataatccta taatattcgc ctccgatagt tttcttcaac ttacggagta ctccagagag
     5641 gagatactgg gaaggaattg cagattcctc caaggtccag aaactgatag agctactgtg
     5701 aggaagatac gggatgccat tgacaatcaa actgaagtga cagtgcaact tatcaattac
     5761 accaagtcag ggaaaaaatt ctggaacctg ttccatttgc aacccatgag ggaccaaaag
     5821 ggcgatgtgc aatatttcat aggcgttcaa cttgatggta ctgaacacgt cagagatgcg
     5881 gcggagcgag aagcagtgat gctgattaag aaaacagcag aggagatcga cgaagcggcg
     5941 aaagagttgt acgaagatga ggaagagttc tacaatgata tgcaaactat cacactgaaa
     6001 tctgagctga actagctgaa gttcatctgc accaaagctt aagtttaaac cgctgatcag
     6061 cctcgactgt gccttctagt tgccagccat ctgttgtttg cccctccccc gtgccttcct
     6121 tgaccctgga aggtgccact cccactgtcc tttcctaata aaatgaggaa attgcatcgc
     6181 attgtctgag taggtgtcat tctattctgg ggggtggggt ggggcaggac agcaaggggg
     6241 aggattggga agacaatagc aggcatgctg gggatgcggt gggctctatg gcttctgagg
     6301 cggaaagaac cagctggggc tctagggggt atccccacgc gccctgtagc ggcgcattaa
     6361 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     6421 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     6481 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     6541 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     6601 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     6661 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     6721 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc tgtggaatgt
     6781 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     6841 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     6901 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     6961 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     7021 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     7081 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     7141 atctgatcaa gagacaggat gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc
     7201 aggttctccg gccgcttggg tggagaggct attcggctat gactgggcac aacagacaat
     7261 cggctgctct gatgccgccg tgttccggct gtcagcgcag gggcgcccgg ttctttttgt
     7321 caagaccgac ctgtccggtg ccctgaatga actgcaggac gaggcagcgc ggctatcgtg
     7381 gctggccacg acgggcgttc cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag
     7441 ggactggctg ctattgggcg aagtgccggg gcaggatctc ctgtcatctc accttgctcc
     7501 tgccgagaaa gtatccatca tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc
     7561 tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga
     7621 agccggtctt gtcgatcagg atgatctgga cgaagagcat caggggctcg cgccagccga
     7681 actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag gatctcgtcg tgacccatgg
     7741 cgatgcctgc ttgccgaata tcatggtgga aaatggccgc ttttctggat tcatcgactg
     7801 tggccggctg ggtgtggcgg accgctatca ggacatagcg ttggctaccc gtgatattgc
     7861 tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg ctttacggta tcgccgctcc
     7921 cgattcgcag cgcatcgcct tctatcgcct tcttgacgag ttcttctgag cgggactctg
     7981 gggttcgaaa tgaccgacca agcgacgccc aacctgccat cacgagattt cgattccacc
     8041 gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg ctggatgatc
     8101 ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt tattgcagct
     8161 tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc atttttttca
     8221 ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt ctgtataccg
     8281 tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt
     8341 tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa agcctggggt
     8401 gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc tttccagtcg
     8461 ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag aggcggtttg
     8521 cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt cgttcggctg
     8581 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     8641 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     8701 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     8761 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     8821 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     8881 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     8941 taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc
     9001 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     9061 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     9121 ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat ctgcgctctg
     9181 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     9241 gctggtagcg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa
     9301 gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa
     9361 gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa
     9421 tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc
     9481 ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga
     9541 ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca
     9601 atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc
     9661 ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat
     9721 tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc
     9781 attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt
     9841 tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc
     9901 ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg
     9961 gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt
    10021 gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg
    10081 gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga
    10141 aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg
    10201 taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg
    10261 tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt
    10321 tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc
    10381 atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca
    10441 tttccccgaa aagtgccacc tgacgtc
//
