LOCUS       sequence-217337-       14237 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 14237)
  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 14237)
  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..14237
                     /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..2658
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          1704..2650
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     2607..2627
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             2679..4529
                     /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      4293..4302
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4563..4610
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(4611..4634,4635..4637,4638..7193,7194..7196,
                     7197..8711)
                     /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             8766..8786
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8811..8960
                     /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             8991..9044
                     /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      9051..9060
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             9060..9446
                     /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             9501..9521
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             9537..10079
                     /codon_start=1
                     /product="C-terminal portion of the p65 subunit of mouse 
                     NF-kappa-B"
                     /label=p65
                     /translation="PSGQISNQALALAPSSAPVLAQTMVPSSAMVPLAQPPAPAPVLTP
                     GPPQSLSAPVPKSTQAGEGTLSEALLHLQFDADEDLGALLGNSTDPGVFTDLASVDNSE
                     FQQLLNQGVSMSHSTAEPMLMEYPEAITRLVTGSQRPPDPAPTPLGTSGLPNGLSGDED
                     FSSIADMDFSALLSQISS"
     CDS             10104..10475
                     /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(10500..11073,11074..11076,11077..11085)
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(10667..10684)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     10894..10913
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             11086..11481
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    11485..11709
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(11730..11754)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     repeat_region   11936..11998
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(12049..12069)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(12049..12067)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(12088..12104)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12088..12104)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12101..12123)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12112..12128
                     /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(12136..12142,12143..12160,12161..12166))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12181..12202
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12319..12336)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12490..13078)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12570..12589)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13249..14040,14041..14109))
                     /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     13872..13891
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14110..14214)
                     /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 agagaaggtg 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 gcctggcttg ggcgctgggg ccgccgcgtg
     1921 cgaatctggt ggcaccttcg cgcctgtctc gctgctttcg ataagtctct agccatttaa
     1981 aatttttgat gacctgctgc gacgcttttt ttctggcaag atagtcttgt aaatgcgggc
     2041 caagatcgat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg acggggcccg
     2101 tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac cgagaatcgg
     2161 acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc cgccgtgtat
     2221 cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag cggaaagatg
     2281 gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct cgggagagcg
     2341 ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg tcgcttcatg
     2401 tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga gcttttggag
     2461 tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc acactgagtg
     2521 ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg aatttgccct
     2581 ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa gtttttttct
     2641 tccatttcag gtgtcgtggg aagcttgggg ccaccatgat tttcaaacca gaagaactac
     2701 gacaggcact gatgccaact ttggaggcac tttaccgtca ggatccagaa tcccttccct
     2761 ttcgtcaacc tgtggaccct cagcttttag gaatccctga ttactttgat attgtgaaga
     2821 gccccatgga tctttctacc attaagagga agttagacac tggacagtat caggagccct
     2881 ggcagtatgt cgatgatatt tggcttatgt tcaataatgc ctggttatat aaccggaaaa
     2941 catcacgggt atacaaatac tgctccaagc tctctgaggt ctttgaacaa gaaattgacc
     3001 cagtgatgca aagccttgga tactgttgtg gcagaaagtt ggagttctct ccacagacac
     3061 tgtgttgcta cggcaaacag ttgtgcacaa tacctcgtga tgccacttat tacagttacc
     3121 agaacaggta tcatttctgt gagaagtgtt tcaatgagat ccaaggggag agcgtttctt
     3181 tgggggatga cccttcccag cctcaaacta caataaataa agaacaattt tccaagagaa
     3241 aaaatgacac actggatcct gaactgtttg ttgaatgtac agagtgcgga agaaagatgc
     3301 atcagatctg tgtccttcac catgagatca tctggcctgc tggattcgtc tgtgatggct
     3361 gtttaaagaa aagtgcacga actaggaaag aaaataagtt ttctgctaaa aggttgccat
     3421 ctaccagact tggcaccttt ctagagaatc gtgtgaatga ctttctgagg cgacagaatc
     3481 accctgagtc aggagaggtc actgttagag tagttcatgc ttctgacaaa accgtggaag
     3541 taaaaccagg catgaaagca aggtttgtgg acagtggaga gatggcagaa tcctttccat
     3601 accgaaccaa agccctcttt gcctttgaag aaattgatgg tgttgacctg tgcttctttg
     3661 gcatgcatgt tcaagagtat ggctctgact gccctccacc caaccagagg agagtataca
     3721 tatcttacct cgatagtgtt catttcttcc gtcctaaatg cttgaggact gcagtctatc
     3781 atgaaatcct aattggatat ttagaatatg tcaagaaatt aggttacaca acagggcata
     3841 tttgggcatg tccaccaagt gagggagatg attatatctt ccattgccat cctcctgacc
     3901 agaagatacc caagcccaag cgactgcagg aatggtacaa aaaaatgctt gacaaggctg
     3961 tatcagagcg tattgtccat gactacaagg atatttttaa acaagctact gaagatagat
     4021 taacaagtgc aaaggaattg ccttatttcg agggtgattt ctggcccaat gttctggaag
     4081 aaagcattaa ggaactggaa caggaggaag aagagagaaa acgagaggaa aacaccagca
     4141 atgaaagcac agatgtgacc aagggagaca gcaaaaatgc taaaaagaag aataataaga
     4201 aaaccagcaa aaataagagc agcctgagta ggggcaacaa gaagaaaccc gggatgccca
     4261 atgtatctaa cgacctctca cagaaactat atgccaccat ggagaagcat aaagaggtct
     4321 tctttgtgat ccgcctcatt gctggccctg ctgccaactc cctgcctccc attgttgatc
     4381 ctgatcctct catcccctgc gatctgatgg atggtcggga tgcgtttctc acgctggcaa
     4441 gggacaagca cctggagttc tcttcactcc gaagagccca gtggtccacc atgtgcatgc
     4501 tggtggagct gcacacgcag agccaggacg gaggtggaag cggaggatgt acggccacca
     4561 tgaaaaggcc ggcggccacg aaaaaggccg gccaggcaaa aaagaaaaag gacaagaagt
     4621 acagcatcgg cctggccatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt
     4681 acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga
     4741 agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc acccggctga
     4801 agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga
     4861 tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct
     4921 tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg
     4981 aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca
     5041 gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc
     5101 ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt
     5161 tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg
     5221 gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc
     5281 tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggcaacctg attgccctga
     5341 gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc
     5401 agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc
     5461 agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca
     5521 tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat
     5581 acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg
     5641 agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacattgacg
     5701 gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg
     5761 gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct
     5821 tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc
     5881 ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga
     5941 ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga
     6001 tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg
     6061 gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg
     6121 agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat aacgagctga
     6181 ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga
     6241 aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga
     6301 aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag
     6361 atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg
     6421 acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac
     6481 tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg
     6541 acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga
     6601 agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt
     6661 ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta
     6721 aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg
     6781 ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg
     6841 acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca
     6901 gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg
     6961 aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc
     7021 agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg
     7081 accaggaact ggacatcaac cggctgtccg actacgatgt ggaccacatc gtgcctcaga
     7141 gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac aaggcccggg
     7201 gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcggc
     7261 agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc aaggccgaga
     7321 gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc
     7381 ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg
     7441 agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg
     7501 atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc
     7561 acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg
     7621 aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga
     7681 gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact
     7741 ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga
     7801 caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc accgtgcgga
     7861 aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct
     7921 tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc agaaagaagg
     7981 actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg
     8041 tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg
     8101 ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca
     8161 agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg
     8221 agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg
     8281 aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc
     8341 tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaagcact
     8401 acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg
     8461 ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc atcagagagc
     8521 aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca
     8581 agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg
     8641 ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc
     8701 tgggaggcga cagcgctgga ggaggtggaa gcggaggagg aggaagcgga ggaggaggta
     8761 gcggacctaa gaaaaagagg aaggtggcgg ccgctggatc cggacgggct gacgcattgg
     8821 acgattttga tctggatatg ctgggaagtg acgccctcga tgattttgac cttgacatgc
     8881 ttggttcgga tgcccttgat gactttgacc tcgacatgct cggcagtgac gcccttgatg
     8941 atttcgacct ggacatgctg attaactgta cgggcggcgg gtccggagga gagggcagag
     9001 gaagtctcct aacatgcggt gacgtggagg agaatcctgg cccatgtacg gccaccatgg
     9061 cttcaaactt tactcagttc gtgctcgtgg acaatggtgg gacaggggat gtgacagtgg
     9121 ctccttctaa tttcgctaat ggggtggcag agtggatcag ctccaactca cggagccagg
     9181 cctacaaggt gacatgcagc gtcaggcagt ctagtgccca gaagagaaag tataccatca
     9241 aggtggaggt ccccaaagtg gctacccaga cagtgggcgg agtcgaactg cctgtcgccg
     9301 cttggaggtc ctacctgaac atggagctca ctatcccaat tttcgctacc aattctgact
     9361 gtgaactcat cgtgaaggca atgcaggggc tcctcaaaga cggtaatcct atcccttccg
     9421 ccatcgccgc taactcaggt atctacagcg ctggaggagg tggaagcgga ggaggaggaa
     9481 gcggaggagg aggtagcgga cctaagaaaa agaggaaggt ggcggccgct ggatcccctt
     9541 cagggcagat cagcaaccag gccctggctc tggcccctag ctccgctcca gtgctggccc
     9601 agactatggt gccctctagt gctatggtgc ctctggccca gccacctgct ccagcccctg
     9661 tgctgacccc aggaccaccc cagtcactga gcgctccagt gcccaagtct acacaggccg
     9721 gcgaggggac tctgagtgaa gctctgctgc acctgcagtt cgacgctgat gaggacctgg
     9781 gagctctgct ggggaacagc accgatcccg gagtgttcac agatctggcc tccgtggaca
     9841 actctgagtt tcagcagctg ctgaatcagg gcgtgtccat gtctcatagt acagccgaac
     9901 caatgctgat ggagtacccc gaagccatta cccggctggt gaccggcagc cagcggcccc
     9961 ccgaccccgc tccaactccc ctgggaacca gcggcctgcc taatgggctg tccggagatg
    10021 aagacttctc aagcatcgct gatatggact ttagtgccct gctgtcacag atttcctcta
    10081 gtgggcaggg aggaggtgga agcggcttca gcgtggacac cagtgccctg ctggacctgt
    10141 tcagcccctc ggtgaccgtg cccgacatga gcctgcctga ccttgacagc agcctggcca
    10201 gtatccaaga gctcctgtct ccccaggagc cccccaggcc tcccgaggca gagaacagca
    10261 gcccggattc agggaagcag ctggtgcact acacagcgca gccgctgttc ctgctggacc
    10321 ccggctccgt ggacaccggg agcaacgacc tgccggtgct gtttgagctg ggagagggct
    10381 cctacttctc cgaaggggac ggcttcgccg aggaccccac catctccctg ctgacaggct
    10441 cggagcctcc caaagccaag gaccccactg tctcctgtac atgagaattc cgcccccccc
    10501 ccctctccct cccccccccc taacgttact ggccgaagcc gcttggaata aggccggtgt
    10561 gcgtttgtct atatgttatt ttccaccata ttgccgtctt ttggcaatgt gagggcccgg
    10621 aaacctggcc ctgtcttctt gacgagcatt cctaggggtc tttcccctct cgccaaagga
    10681 atgcaaggtc tgttgaatgt cgtgaaggaa gcagttcctc tggaagcttc ttgaagacaa
    10741 acaacgtctg tagcgaccct ttgcaggcag cggaaccccc cacctggcga caggtgcctc
    10801 tgcggccaaa agccacgtgt ataagataca cctgcaaagg cggcacaacc ccagtgccac
    10861 gttgtgagtt ggatagttgt ggaaagagtc aaatggctct cctcaagcgt attcaacaag
    10921 gggctgaagg atgcccagaa ggtaccccat tgtatgggat ctgatctggg gcctcggtgc
    10981 acatgcttta catgtgttta gtcgaggtta aaaaacgtct aggccccccg aaccacgggg
    11041 acgtggtttt cctttgaaaa acacgatgat aatatggcca caaccatggc caagcctttg
    11101 tctcaagaag aatccaccct cattgaaaga gcaacggcta caatcaacag catccccatc
    11161 tctgaagact acagcgtcgc cagcgcagct ctctctagcg acggccgcat cttcactggt
    11221 gtcaatgtat atcattttac tgggggacct tgtgcagaac tcgtggtgct gggcactgct
    11281 gctgctgcgg cagctggcaa cctgacttgt atcgtcgcga tcggaaatga gaacaggggc
    11341 atcttgagcc cctgcggacg gtgccgacag gtgcttctcg atctgcatcc tgggatcaaa
    11401 gccatagtga aggacagtga tggacagccg acggcagttg ggattcgtga attgctgccc
    11461 tctggttatg tgtgggaggg ccgactgtgc cttctagttg ccagccatct gttgtttgcc
    11521 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
    11581 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
    11641 ggcaggacag caagggggag gattgggaag tcaatagcag gcatgctggg gatgcggtgg
    11701 gctctatgga ctagtctcga gatatctaga cccagctttc ttgtacaaag tggtactagt
    11761 taaaagtttt gttactttat agaagaaatt ttgagttttt gttttttttt aataaataaa
    11821 taaacataaa taaattgttt gttgaattta ttattagtat gtaagtgtaa atataataaa
    11881 acttaatatc tattcaaatt aataaataaa cctcgatata cagaccgata aaacacatgc
    11941 gtcaatttta cgcatgatta tctttaacgt acgtcacaat atgattatct ttctagggtt
    12001 aatctagtat acgcgtatgc ggccgcttaa ttaatccagc ttttgttccc tttagtgagg
    12061 gttaattgcg cgcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
    12121 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
    12181 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
    12241 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
    12301 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
    12361 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
    12421 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
    12481 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
    12541 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
    12601 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
    12661 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
    12721 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
    12781 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
    12841 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
    12901 gtggtggcct aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa
    12961 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
    13021 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
    13081 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
    13141 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
    13201 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
    13261 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
    13321 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
    13381 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
    13441 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
    13501 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
    13561 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
    13621 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
    13681 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
    13741 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
    13801 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
    13861 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
    13921 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
    13981 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
    14041 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
    14101 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
    14161 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
    14221 tccccgaaaa gtgccac
//
