LOCUS       sequence-227309-       14236 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses p300-dCas9-VP64 and MS2-p65-HSF1 fusion proteins.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14236)
  AUTHORS   Chong ZS, Ohnishi S, Yusa K, Wright GJ
  TITLE     Pooled extracellular receptor-ligand interaction screening using 
            CRISPR activation.
  JOURNAL   Genome Biol. 2018 Nov 26;19(1):205. doi: 10.1186/s13059-018-1581-3.
  PUBMED    30477585
REFERENCE   2  (bases 1 to 14236)
  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: "Genome 
            Biol."; date: "2018-11-26"; volume: "19(1):205. doi"; pages: " 
            10.1186/s13059-018-1581-3"
COMMENT     Sequence Label: pPB-R1R2_EF1ap300dCas9VP64_T2A_MS2p65HSF1-IRES...
FEATURES             Location/Qualifiers
     source          1..14236
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(30..458)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(140..161)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     352..371
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     585..607
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     599..616
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     626..645
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     repeat_region   674..708
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    998..1022
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     promoter        1473..2654
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1704..2646
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2603..2623
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             2674..4524
                     /codon_start=1
                     /gene="EP300"
                     /product="catalytic histone acetyltransferase core of the 
                     human E1A-associated protein p300 (Delvecchio et al., 
                     2013)"
                     /label=p300 core
                     /translation="IFKPEELRQALMPTLEALYRQDPESLPFRQPVDPQLLGIPDYFDI
                     VKSPMDLSTIKRKLDTGQYQEPWQYVDDIWLMFNNAWLYNRKTSRVYKYCSKLSEVFEQ
                     EIDPVMQSLGYCCGRKLEFSPQTLCCYGKQLCTIPRDATYYSYQNRYHFCEKCFNEIQG
                     ESVSLGDDPSQPQTTINKEQFSKRKNDTLDPELFVECTECGRKMHQICVLHHEIIWPAG
                     FVCDGCLKKSARTRKENKFSAKRLPSTRLGTFLENRVNDFLRRQNHPESGEVTVRVVHA
                     SDKTVEVKPGMKARFVDSGEMAESFPYRTKALFAFEEIDGVDLCFFGMHVQEYGSDCPP
                     PNQRRVYISYLDSVHFFRPKCLRTAVYHEILIGYLEYVKKLGYTTGHIWACPPSEGDDY
                     IFHCHPPDQKIPKPKRLQEWYKKMLDKAVSERIVHDYKDIFKQATEDRLTSAKELPYFE
                     GDFWPNVLEESIKELEQEEEERKREENTSNESTDVTKGDSKNAKKKNNKKTSKNKSSLS
                     RGNKKKPGMPNVSNDLSQKLYATMEKHKEVFFVIRLIAGPAANSLPPIVDPDPLIPCDL
                     MDGRDAFLTLARDKHLEFSSLRRAQWSTMCMLVELHTQSQD"
     regulatory      4288..4297
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4558..4605
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(4606..4629,4630..4632,4633..7188,7189..7191,
                     7192..8706)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the N863A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKARGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             8761..8781
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8806..8955
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             8986..9039
                     /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"
     regulatory      9046..9055
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             9055..9441
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             9496..9516
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             9532..10074
                     /codon_start=1
                     /product="C-terminal portion of the p65 subunit of mouse 
                     NF-kappa-B"
                     /label=p65
                     /translation="PSGQISNQALALAPSSAPVLAQTMVPSSAMVPLAQPPAPAPVLTP
                     GPPQSLSAPVPKSTQAGEGTLSEALLHLQFDADEDLGALLGNSTDPGVFTDLASVDNSE
                     FQQLLNQGVSMSHSTAEPMLMEYPEAITRLVTGSQRPPDPAPTPLGTSGLPNGLSGDED
                     FSSIADMDFSALLSQISS"
     CDS             10099..10470
                     /codon_start=1
                     /product="C-terminal activation domain from the human heat 
                     shock transcription factor HSF1"
                     /label=HSF1
                     /translation="GFSVDTSALLDLFSPSVTVPDMSLPDLDSSLASIQELLSPQEPPR
                     PPEAENSSPDSGKQLVHYTAQPLFLLDPGSVDTGSNDLPVLFELGEGSYFSEGDGFAED
                     PTISLLTGSEPPKAKDPTVS"
     misc_feature    join(10495..11069,11070..11072,11073..11081)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(10662..10679)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     10889..10908
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             11082..11480
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    11484..11708
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(11729..11753)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     repeat_region   11935..11997
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(12048..12068)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(12048..12066)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(12087..12103)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12087..12103)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12100..12122)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12111..12127
                     /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(12135..12141,12142..12159,12160..12165))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12180..12201
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12318..12335)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12489..13077)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12569..12588)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13248..14039,14040..14108))
                     /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     13871..13890
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14109..14213)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
       61 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
      121 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
      181 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
      241 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
      301 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
      361 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
      421 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctgcg
      481 caactgttgg gaagggcgat cggtgcgggc ctcttcgcta ttacgccagc tggcgaaagg
      541 gggatgtgct gcaaggcgat taagttgggt aacgccaggg ttttcccagt cacgacgttg
      601 taaaacgacg gccagtgagc gcgcgtaata cgactcacta tagggcgaat tggggcgcgc
      661 cattctagat taaccctaga aagatagtct gcgtaaaatt gacgcatgca ttcttgaaat
      721 attgctctct ctttctaaat agcgcgaatc cgtcgctgtg catttaggac atctcagtcg
      781 ccgcttggag ctcccgtgag gcgtgcttgt caatgcggta agtgtcactg attttgaact
      841 ataacgaccg cgtgagtcaa aatgacgcat gattatcttt tacgtgactt ttaagattta
      901 actcatacga taattatatt gttatttcat gttctactta cgtgataact tattatatat
      961 atattttctt gttatagata tcaactagaa tgctagcaca agtttgtaca aaaaagcagg
     1021 ctggcgccgg aaccaattca gtcgactgga tccacgcgtg aggggcgcgg aagccggggc
     1081 ggaggaggag ccaaccccgg gctgggctga gacccgcaga ggaagacgct ctagggattt
     1141 gtcccggact agcgagatgg caaggctgag gacgggaggc tgattgagag gcgaaggtac
     1201 accctaatct caatacaacc tttggagcta agccagcaat ggtagaggga agattctgca
     1261 cgtcccttcc aggcggcctc cccgtcacca ccccccccaa cccgccccga ccggagctga
     1321 gagtaattca tacaaaagga ctcgcccctg ccttggggaa tcccagggac cgtcgttaaa
     1381 ctcccactaa cgtagaaccc agagatcgct gcgttcccgc cccctcaccc gcccgctctc
     1441 gtcatcactg aggtggagaa gagcatgcgt gaggctccgg tgcccgtcag tgggcagagc
     1501 gcacatcgcc cacagtcccc gagaagttgg ggggaggggt cggcaattga accggtgcct
     1561 agagaaagtg gcgcggggta aactgggaaa gtgatgtcgt gtactggctc cgcctttttc
     1621 ccgagggtgg gggagaaccg tatataagtg cagtagtcgc cgtgaacgtt ctttttcgca
     1681 acgggtttgc cgccagaaca caggtaagtg ccgtgtgtgg ttcccgcggg cctggcctct
     1741 ttacgggtta tggcccttgc gtgccttgaa ttacttccac gcccctggct gcagtacgtg
     1801 attcttgatc ccgagcttcg ggttggaagt gggtgggaga gttcgaggcc ttgcgcttaa
     1861 ggagcccctt cgcctcgtgc ttgagttgag gcctggcctg ggcgctgggg ccgccgcgtg
     1921 cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct agccatttaa
     1981 aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt aaatgcgggc
     2041 caagatctgc acactggtat ttcggttttt ggggccgcgg gcggcgacgg ggcccgtgcg
     2101 tcccagcgca catgttcggc gaggcggggc ctgcgagcgc ggccaccgag aatcggacgg
     2161 gggtagtctc aagctggccg gcctgctctg gtgcctggcc tcgcgccgcc gtgtatcgcc
     2221 ccgccctggg cggcaaggct ggcccggtcg gcaccagttg cgtgagcgga aagatggccg
     2281 cttcccggcc ctgctgcagg gagctcaaaa tggaggacgc ggcgctcggg agagcgggcg
     2341 ggtgagtcac ccacacaaag gaaaagggcc tttccgtcct cagccgtcgc ttcatgtgac
     2401 tccacggagt accgggcgcc gtccaggcac ctcgattagt tctcgagctt ttggagtacg
     2461 tcgtctttag gttgggggga ggggttttat gcgatggagt ttccccacac tgagtgggtg
     2521 gagactgaag ttaggccagc ttggcacttg atgtaattct ccttggaatt tgcccttttt
     2581 gagtttggat cttggttcat tctcaagcct cagacagtgg ttcaaagttt ttttcttcca
     2641 tttcaggtgt cgtggaagct tggggccacc atgattttca aaccagaaga actacgacag
     2701 gcactgatgc caactttgga ggcactttac cgtcaggatc cagaatccct tccctttcgt
     2761 caacctgtgg accctcagct tttaggaatc cctgattact ttgatattgt gaagagcccc
     2821 atggatcttt ctaccattaa gaggaagtta gacactggac agtatcagga gccctggcag
     2881 tatgtcgatg atatttggct tatgttcaat aatgcctggt tatataaccg gaaaacatca
     2941 cgggtataca aatactgctc caagctctct gaggtctttg aacaagaaat tgacccagtg
     3001 atgcaaagcc ttggatactg ttgtggcaga aagttggagt tctctccaca gacactgtgt
     3061 tgctacggca aacagttgtg cacaatacct cgtgatgcca cttattacag ttaccagaac
     3121 aggtatcatt tctgtgagaa gtgtttcaat gagatccaag gggagagcgt ttctttgggg
     3181 gatgaccctt cccagcctca aactacaata aataaagaac aattttccaa gagaaaaaat
     3241 gacacactgg atcctgaact gtttgttgaa tgtacagagt gcggaagaaa gatgcatcag
     3301 atctgtgtcc ttcaccatga gatcatctgg cctgctggat tcgtctgtga tggctgttta
     3361 aagaaaagtg cacgaactag gaaagaaaat aagttttctg ctaaaaggtt gccatctacc
     3421 agacttggca cctttctaga gaatcgtgtg aatgactttc tgaggcgaca gaatcaccct
     3481 gagtcaggag aggtcactgt tagagtagtt catgcttctg acaaaaccgt ggaagtaaaa
     3541 ccaggcatga aagcaaggtt tgtggacagt ggagagatgg cagaatcctt tccataccga
     3601 accaaagccc tctttgcctt tgaagaaatt gatggtgttg acctgtgctt ctttggcatg
     3661 catgttcaag agtatggctc tgactgccct ccacccaacc agaggagagt atacatatct
     3721 tacctcgata gtgttcattt cttccgtcct aaatgcttga ggactgcagt ctatcatgaa
     3781 atcctaattg gatatttaga atatgtcaag aaattaggtt acacaacagg gcatatttgg
     3841 gcatgtccac caagtgaggg agatgattat atcttccatt gccatcctcc tgaccagaag
     3901 atacccaagc ccaagcgact gcaggaatgg tacaaaaaaa tgcttgacaa ggctgtatca
     3961 gagcgtattg tccatgacta caaggatatt tttaaacaag ctactgaaga tagattaaca
     4021 agtgcaaagg aattgcctta tttcgagggt gatttctggc ccaatgttct ggaagaaagc
     4081 attaaggaac tggaacagga ggaagaagag agaaaacgag aggaaaacac cagcaatgaa
     4141 agcacagatg tgaccaaggg agacagcaaa aatgctaaaa agaagaataa taagaaaacc
     4201 agcaaaaata agagcagcct gagtaggggc aacaagaaga aacccgggat gcccaatgta
     4261 tctaacgacc tctcacagaa actatatgcc accatggaga agcataaaga ggtcttcttt
     4321 gtgatccgcc tcattgctgg ccctgctgcc aactccctgc ctcccattgt tgatcctgat
     4381 cctctcatcc cctgcgatct gatggatggt cgggatgcgt ttctcacgct ggcaagggac
     4441 aagcacctgg agttctcttc actccgaaga gcccagtggt ccaccatgtg catgctggtg
     4501 gagctgcaca cgcagagcca ggacggaggt ggaagcggag gatgtacggc caccatgaaa
     4561 aggccggcgg ccacgaaaaa ggccggccag gcaaaaaaga aaaaggacaa gaagtacagc
     4621 atcggcctgg ccatcggcac caactctgtg ggctgggccg tgatcaccga cgagtacaag
     4681 gtgcccagca agaaattcaa ggtgctgggc aacaccgacc ggcacagcat caagaagaac
     4741 ctgatcggag ccctgctgtt cgacagcggc gaaacagccg aggccacccg gctgaagaga
     4801 accgccagaa gaagatacac cagacggaag aaccggatct gctatctgca agagatcttc
     4861 agcaacgaga tggccaaggt ggacgacagc ttcttccaca gactggaaga gtccttcctg
     4921 gtggaagagg ataagaagca cgagcggcac cccatcttcg gcaacatcgt ggacgaggtg
     4981 gcctaccacg agaagtaccc caccatctac cacctgagaa agaaactggt ggacagcacc
     5041 gacaaggccg acctgcggct gatctatctg gccctggccc acatgatcaa gttccggggc
     5101 cacttcctga tcgagggcga cctgaacccc gacaacagcg acgtggacaa gctgttcatc
     5161 cagctggtgc agacctacaa ccagctgttc gaggaaaacc ccatcaacgc cagcggcgtg
     5221 gacgccaagg ccatcctgtc tgccagactg agcaagagca gacggctgga aaatctgatc
     5281 gcccagctgc ccggcgagaa gaagaatggc ctgttcggca acctgattgc cctgagcctg
     5341 ggcctgaccc ccaacttcaa gagcaacttc gacctggccg aggatgccaa actgcagctg
     5401 agcaaggaca cctacgacga cgacctggac aacctgctgg cccagatcgg cgaccagtac
     5461 gccgacctgt ttctggccgc caagaacctg tccgacgcca tcctgctgag cgacatcctg
     5521 agagtgaaca ccgagatcac caaggccccc ctgagcgcct ctatgatcaa gagatacgac
     5581 gagcaccacc aggacctgac cctgctgaaa gctctcgtgc ggcagcagct gcctgagaag
     5641 tacaaagaga ttttcttcga ccagagcaag aacggctacg ccggctacat tgacggcgga
     5701 gccagccagg aagagttcta caagttcatc aagcccatcc tggaaaagat ggacggcacc
     5761 gaggaactgc tcgtgaagct gaacagagag gacctgctgc ggaagcagcg gaccttcgac
     5821 aacggcagca tcccccacca gatccacctg ggagagctgc acgccattct gcggcggcag
     5881 gaagattttt acccattcct gaaggacaac cgggaaaaga tcgagaagat cctgaccttc
     5941 cgcatcccct actacgtggg ccctctggcc aggggaaaca gcagattcgc ctggatgacc
     6001 agaaagagcg aggaaaccat caccccctgg aacttcgagg aagtggtgga caagggcgct
     6061 tccgcccaga gcttcatcga gcggatgacc aacttcgata agaacctgcc caacgagaag
     6121 gtgctgccca agcacagcct gctgtacgag tacttcaccg tgtataacga gctgaccaaa
     6181 gtgaaatacg tgaccgaggg aatgagaaag cccgccttcc tgagcggcga gcagaaaaag
     6241 gccatcgtgg acctgctgtt caagaccaac cggaaagtga ccgtgaagca gctgaaagag
     6301 gactacttca agaaaatcga gtgcttcgac tccgtggaaa tctccggcgt ggaagatcgg
     6361 ttcaacgcct ccctgggcac ataccacgat ctgctgaaaa ttatcaagga caaggacttc
     6421 ctggacaatg aggaaaacga ggacattctg gaagatatcg tgctgaccct gacactgttt
     6481 gaggacagag agatgatcga ggaacggctg aaaacctatg cccacctgtt cgacgacaaa
     6541 gtgatgaagc agctgaagcg gcggagatac accggctggg gcaggctgag ccggaagctg
     6601 atcaacggca tccgggacaa gcagtccggc aagacaatcc tggatttcct gaagtccgac
     6661 ggcttcgcca acagaaactt catgcagctg atccacgacg acagcctgac ctttaaagag
     6721 gacatccaga aagcccaggt gtccggccag ggcgatagcc tgcacgagca cattgccaat
     6781 ctggccggca gccccgccat taagaagggc atcctgcaga cagtgaaggt ggtggacgag
     6841 ctcgtgaaag tgatgggccg gcacaagccc gagaacatcg tgatcgaaat ggccagagag
     6901 aaccagacca cccagaaggg acagaagaac agccgcgaga gaatgaagcg gatcgaagag
     6961 ggcatcaaag agctgggcag ccagatcctg aaagaacacc ccgtggaaaa cacccagctg
     7021 cagaacgaga agctgtacct gtactacctg cagaatgggc gggatatgta cgtggaccag
     7081 gaactggaca tcaaccggct gtccgactac gatgtggacc acatcgtgcc tcagagcttt
     7141 ctgaaggacg actccatcga caacaaggtg ctgaccagaa gcgacaaggc ccggggcaag
     7201 agcgacaacg tgccctccga agaggtcgtg aagaagatga agaactactg gcggcagctg
     7261 ctgaacgcca agctgattac ccagagaaag ttcgacaatc tgaccaaggc cgagagaggc
     7321 ggcctgagcg aactggataa ggccggcttc atcaagagac agctggtgga aacccggcag
     7381 atcacaaagc acgtggcaca gatcctggac tcccggatga acactaagta cgacgagaat
     7441 gacaagctga tccgggaagt gaaagtgatc accctgaagt ccaagctggt gtccgatttc
     7501 cggaaggatt tccagtttta caaagtgcgc gagatcaaca actaccacca cgcccacgac
     7561 gcctacctga acgccgtcgt gggaaccgcc ctgatcaaaa agtaccctaa gctggaaagc
     7621 gagttcgtgt acggcgacta caaggtgtac gacgtgcgga agatgatcgc caagagcgag
     7681 caggaaatcg gcaaggctac cgccaagtac ttcttctaca gcaacatcat gaactttttc
     7741 aagaccgaga ttaccctggc caacggcgag atccggaagc ggcctctgat cgagacaaac
     7801 ggcgaaaccg gggagatcgt gtgggataag ggccgggatt ttgccaccgt gcggaaagtg
     7861 ctgagcatgc cccaagtgaa tatcgtgaaa aagaccgagg tgcagacagg cggcttcagc
     7921 aaagagtcta tcctgcccaa gaggaacagc gataagctga tcgccagaaa gaaggactgg
     7981 gaccctaaga agtacggcgg cttcgacagc cccaccgtgg cctattctgt gctggtggtg
     8041 gccaaagtgg aaaagggcaa gtccaagaaa ctgaagagtg tgaaagagct gctggggatc
     8101 accatcatgg aaagaagcag cttcgagaag aatcccatcg actttctgga agccaagggc
     8161 tacaaagaag tgaaaaagga cctgatcatc aagctgccta agtactccct gttcgagctg
     8221 gaaaacggcc ggaagagaat gctggcctct gccggcgaac tgcagaaggg aaacgaactg
     8281 gccctgccct ccaaatatgt gaacttcctg tacctggcca gccactatga gaagctgaag
     8341 ggctcccccg aggataatga gcagaaacag ctgtttgtgg aacagcacaa gcactacctg
     8401 gacgagatca tcgagcagat cagcgagttc tccaagagag tgatcctggc cgacgctaat
     8461 ctggacaaag tgctgtccgc ctacaacaag caccgggata agcccatcag agagcaggcc
     8521 gagaatatca tccacctgtt taccctgacc aatctgggag cccctgccgc cttcaagtac
     8581 tttgacacca ccatcgaccg gaagaggtac accagcacca aagaggtgct ggacgccacc
     8641 ctgatccacc agagcatcac cggcctgtac gagacacgga tcgacctgtc tcagctggga
     8701 ggcgacagcg ctggaggagg tggaagcgga ggaggaggaa gcggaggagg aggtagcgga
     8761 cctaagaaaa agaggaaggt ggcggccgct ggatccggac gggctgacgc attggacgat
     8821 tttgatctgg atatgctggg aagtgacgcc ctcgatgatt ttgaccttga catgcttggt
     8881 tcggatgccc ttgatgactt tgacctcgac atgctcggca gtgacgccct tgatgatttc
     8941 gacctggaca tgctgattaa ctgtacgggc ggcgggtccg gaggagaggg cagaggaagt
     9001 ctcctaacat gcggtgacgt ggaggagaat cctggcccat gtacggccac catggcttca
     9061 aactttactc agttcgtgct cgtggacaat ggtgggacag gggatgtgac agtggctcct
     9121 tctaatttcg ctaatggggt ggcagagtgg atcagctcca actcacggag ccaggcctac
     9181 aaggtgacat gcagcgtcag gcagtctagt gcccagaaga gaaagtatac catcaaggtg
     9241 gaggtcccca aagtggctac ccagacagtg ggcggagtcg aactgcctgt cgccgcttgg
     9301 aggtcctacc tgaacatgga gctcactatc ccaattttcg ctaccaattc tgactgtgaa
     9361 ctcatcgtga aggcaatgca ggggctcctc aaagacggta atcctatccc ttccgccatc
     9421 gccgctaact caggtatcta cagcgctgga ggaggtggaa gcggaggagg aggaagcgga
     9481 ggaggaggta gcggacctaa gaaaaagagg aaggtggcgg ccgctggatc cccttcaggg
     9541 cagatcagca accaggccct ggctctggcc cctagctccg ctccagtgct ggcccagact
     9601 atggtgccct ctagtgctat ggtgcctctg gcccagccac ctgctccagc ccctgtgctg
     9661 accccaggac caccccagtc actgagcgct ccagtgccca agtctacaca ggccggcgag
     9721 gggactctga gtgaagctct gctgcacctg cagttcgacg ctgatgagga cctgggagct
     9781 ctgctgggga acagcaccga tcccggagtg ttcacagatc tggcctccgt ggacaactct
     9841 gagtttcagc agctgctgaa tcagggcgtg tccatgtctc atagtacagc cgaaccaatg
     9901 ctgatggagt accccgaagc cattacccgg ctggtgaccg gcagccagcg gccccccgac
     9961 cccgctccaa ctcccctggg aaccagcggc ctgcctaatg ggctgtccgg agatgaagac
    10021 ttctcaagca tcgctgatat ggactttagt gccctgctgt cacagatttc ctctagtggg
    10081 cagggaggag gtggaagcgg cttcagcgtg gacaccagtg ccctgctgga cctgttcagc
    10141 ccctcggtga ccgtgcccga catgagcctg cctgaccttg acagcagcct ggccagtatc
    10201 caagagctcc tgtctcccca ggagcccccc aggcctcccg aggcagagaa cagcagcccg
    10261 gattcaggga agcagctggt gcactacaca gcgcagccgc tgttcctgct ggaccccggc
    10321 tccgtggaca ccgggagcaa cgacctgccg gtgctgtttg agctgggaga gggctcctac
    10381 ttctccgaag gggacggctt cgccgaggac cccaccatct ccctgctgac aggctcggag
    10441 cctcccaaag ccaaggaccc cactgtctcc tgtacatgag aattccgccc cccccccctc
    10501 tccctccccc ccccctaacg ttactggccg aagccgcttg gaataaggcc ggtgtgcgtt
    10561 tgtctatatg ttattttcca ccatattgcc gtcttttggc aatgtgaggg cccggaaacc
    10621 tggccctgtc ttcttgacga gcattcctag gggtctttcc cctctcgcca aaggaatgca
    10681 aggtctgttg aatgtcgtga aggaagcagt tcctctggaa gcttcttgaa gacaaacaac
    10741 gtctgtagcg accctttgca ggcagcggaa ccccccacct ggcgacaggt gcctctgcgg
    10801 ccaaaagcca cgtgtataag atacacctgc aaaggcggca caaccccagt gccacgttgt
    10861 gagttggata gttgtggaaa gagtcaaatg gctctcctca agcgtattca acaaggggct
    10921 gaaggatgcc cagaaggtac cccattgtat gggatctgat ctggggcctc ggtgcacatg
    10981 ctttacatgt gtttagtcga ggttaaaaaa acgtctaggc cccccgaacc acggggacgt
    11041 ggttttcctt tgaaaaacac gatgataata tggccacaac catggccaag cctttgtctc
    11101 aagaagaatc caccctcatt gaaagagcaa cggctacaat caacagcatc cccatctctg
    11161 aagactacag cgtcgccagc gcagctctct ctagcgacgg ccgcatcttc actggtgtca
    11221 atgtatatca ttttactggg ggaccttgtg cagaactcgt ggtgctgggc actgctgctg
    11281 ctgcggcagc tggcaacctg acttgtatcg tcgcgatcgg aaatgagaac aggggcatct
    11341 tgagcccctg cggacggtgc cgacaggtgc ttctcgatct gcatcctggg atcaaagcca
    11401 tagtgaagga cagtgatgga cagccgacgg cagttgggat tcgtgaattg ctgccctctg
    11461 gttatgtgtg ggagggctga cgactgtgcc ttctagttgc cagccatctg ttgtttgccc
    11521 ctcccccgtg ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa
    11581 tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg
    11641 gcaggacagc aagggggagg attgggaagt caatagcagg catgctgggg atgcggtggg
    11701 ctctatggac tagtctcgag atatctagac ccagctttct tgtacaaagt ggtactagtt
    11761 aaaagttttg ttactttata gaagaaattt tgagtttttg ttttttttta ataaataaat
    11821 aaacataaat aaattgtttg ttgaatttat tattagtatg taagtgtaaa tataataaaa
    11881 cttaatatct attcaaatta ataaataaac ctcgatatac agaccgataa aacacatgcg
    11941 tcaattttac gcatgattat ctttaacgta cgtcacaata tgattatctt tctagggtta
    12001 atctagtata cgcgtatgcg gccgcttaat taatccagct tttgttccct ttagtgaggg
    12061 ttaattgcgc gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg
    12121 ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg gggtgcctaa
    12181 tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
    12241 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
    12301 gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg gctgcggcga
    12361 gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg ggataacgca
    12421 ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg
    12481 ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt
    12541 cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc
    12601 ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct
    12661 tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc
    12721 gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta
    12781 tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca
    12841 gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag
    12901 tggtggccta actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag
    12961 ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac caccgctggt
    13021 agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa
    13081 gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg
    13141 attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga
    13201 agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta ccaatgctta
    13261 atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt tgcctgactc
    13321 cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag tgctgcaatg
    13381 ataccgcgag acccacgctc accggctcca gatttatcag caataaacca gccagccgga
    13441 agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc tattaattgt
    13501 tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt
    13561 gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag ctccggttcc
    13621 caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt tagctccttc
    13681 ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat ggttatggca
    13741 gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt gactggtgag
    13801 tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc ttgcccggcg
    13861 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
    13921 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
    13981 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
    14041 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
    14101 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
    14161 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
    14221 ccccgaaaag tgccac
//
