LOCUS       sequence-198593-       14555 bp    DNA     linear   UNA 14-APR-2025
DEFINITION  Piggybac transposon DEST vector encoding dCAS9-VP64 and MS2-P65-HSF1
            activator helper complex. It is used for insertion of multiple 
            U6-sgRNA expression cassettes..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 14555)
  AUTHORS   Li S, Zhang A, Xue H, Li D, Liu Y
  TITLE     One-Step piggyBac Transposon-Based CRISPR/Cas9 Activation of 
            Multiple Genes.
  JOURNAL   Mol Ther Nucleic Acids. 2017 Sep 15;8:64-76. doi: 
            10.1016/j.omtn.2017.06.007. Epub 2017 Jun 15.
  PUBMED    28918057
REFERENCE   2  (bases 1 to 14555)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Mol Ther 
            Nucleic Acids."; date: "2017-09-15"; volume: "8:64-76. doi"; pages: 
            " 10.1016/j.omtn.2017.06.007. Epub 2017 Jun 15"
COMMENT     Sequence Label: PB-SAM-Dest
FEATURES             Location/Qualifiers
     source          1..14555
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     primer_bind     74..93
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             126..173
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(174..197,198..200,201..2756,2757..2759,2760..4274)
                     /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             4329..4349
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4374..4523
                     /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             4545..4598
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             4599..4997
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    5139..5260
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     protein_bind    5269..5393
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(5418..5423,5424..5441,5442..5448)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             5502..6161
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(5568..5587)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             6503..6808
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     6687..6706
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(6849..6973)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     repeat_region   7323..7357
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7481..7500)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(7482..7500)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(7587..7604)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7758..8346)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7838..7857)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8517..9308,9309..9377))
                     /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     9140..9159
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(9378..9482)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     9550..9568
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(9606..9628)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     9728..9747
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        9828..9846
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     9828..9845
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    9912..9933
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(9948..9953,9954..9971,9972..9978)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9986..10002
                     /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)."
     primer_bind     9991..10013
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     10010..10026
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     10010..10026
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        10047..10065
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   10248..10310
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     promoter        10613..11791
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          10844..11782
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     11739..11759
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     CDS             11804..12190
                     /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             12245..12265
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             12281..12823
                     /codon_start=1
                     /product="C-terminal portion of the p65 subunit of mouse 
                     NF-kappa-B"
                     /label=p65
                     /translation="PSGQISNQALALAPSSAPVLAQTMVPSSAMVPLAQPPAPAPVLTP
                     GPPQSLSAPVPKSTQAGEGTLSEALLHLQFDADEDLGALLGNSTDPGVFTDLASVDNSE
                     FQQLLNQGVSMSHSTAEPMLMEYPEAITRLVTGSQRPPDPAPTPLGTSGLPNGLSGDED
                     FSSIADMDFSALLSQISS"
     CDS             12848..13219
                     /codon_start=1
                     /product="C-terminal activation domain from the human heat 
                     shock transcription factor HSF1"
                     /label=HSF1
                     /translation="GFSVDTSALLDLFSPSVTVPDMSLPDLDSSLASIQELLSPQEPPR
                     PPEAENSSPDSGKQLVHYTAQPLFLLDPGSVDTGSNDLPVLFELGEGSYFSEGDGFAED
                     PTISLLTGSEPPKAKDPTVS"
     CDS             13235..13288
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             13292..14311
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     "
     polyA_signal    14434..14555
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 tggcgtgtga ccggcggctc tagagcctct gctaaccatg ttcatgcctt cttctttttc
       61 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caaagattta
      121 ccatgaaaag gccggcggcc acgaaaaagg ccggccaggc aaaaaagaaa aaggacaaga
      181 agtacagcat cggcctggcc atcggcacca actctgtggg ctgggccgtg atcaccgacg
      241 agtacaaggt gcccagcaag aaattcaagg tgctgggcaa caccgaccgg cacagcatca
      301 agaagaacct gatcggagcc ctgctgttcg acagcggcga aacagccgag gccacccggc
      361 tgaagagaac cgccagaaga agatacacca gacggaagaa ccggatctgc tatctgcaag
      421 agatcttcag caacgagatg gccaaggtgg acgacagctt cttccacaga ctggaagagt
      481 ccttcctggt ggaagaggat aagaagcacg agcggcaccc catcttcggc aacatcgtgg
      541 acgaggtggc ctaccacgag aagtacccca ccatctacca cctgagaaag aaactggtgg
      601 acagcaccga caaggccgac ctgcggctga tctatctggc cctggcccac atgatcaagt
      661 tccggggcca cttcctgatc gagggcgacc tgaaccccga caacagcgac gtggacaagc
      721 tgttcatcca gctggtgcag acctacaacc agctgttcga ggaaaacccc atcaacgcca
      781 gcggcgtgga cgccaaggcc atcctgtctg ccagactgag caagagcaga cggctggaaa
      841 atctgatcgc ccagctgccc ggcgagaaga agaatggcct gttcggcaac ctgattgccc
      901 tgagcctggg cctgaccccc aacttcaaga gcaacttcga cctggccgag gatgccaaac
      961 tgcagctgag caaggacacc tacgacgacg acctggacaa cctgctggcc cagatcggcg
     1021 accagtacgc cgacctgttt ctggccgcca agaacctgtc cgacgccatc ctgctgagcg
     1081 acatcctgag agtgaacacc gagatcacca aggcccccct gagcgcctct atgatcaaga
     1141 gatacgacga gcaccaccag gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc
     1201 ctgagaagta caaagagatt ttcttcgacc agagcaagaa cggctacgcc ggctacattg
     1261 acggcggagc cagccaggaa gagttctaca agttcatcaa gcccatcctg gaaaagatgg
     1321 acggcaccga ggaactgctc gtgaagctga acagagagga cctgctgcgg aagcagcgga
     1381 ccttcgacaa cggcagcatc ccccaccaga tccacctggg agagctgcac gccattctgc
     1441 ggcggcagga agatttttac ccattcctga aggacaaccg ggaaaagatc gagaagatcc
     1501 tgaccttccg catcccctac tacgtgggcc ctctggccag gggaaacagc agattcgcct
     1561 ggatgaccag aaagagcgag gaaaccatca ccccctggaa cttcgaggaa gtggtggaca
     1621 agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa cttcgataag aacctgccca
     1681 acgagaaggt gctgcccaag cacagcctgc tgtacgagta cttcaccgtg tataacgagc
     1741 tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc cgccttcctg agcggcgagc
     1801 agaaaaaggc catcgtggac ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc
     1861 tgaaagagga ctacttcaag aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg
     1921 aagatcggtt caacgcctcc ctgggcacat accacgatct gctgaaaatt atcaaggaca
     1981 aggacttcct ggacaatgag gaaaacgagg acattctgga agatatcgtg ctgaccctga
     2041 cactgtttga ggacagagag atgatcgagg aacggctgaa aacctatgcc cacctgttcg
     2101 acgacaaagt gatgaagcag ctgaagcggc ggagatacac cggctggggc aggctgagcc
     2161 ggaagctgat caacggcatc cgggacaagc agtccggcaa gacaatcctg gatttcctga
     2221 agtccgacgg cttcgccaac agaaacttca tgcagctgat ccacgacgac agcctgacct
     2281 ttaaagagga catccagaaa gcccaggtgt ccggccaggg cgatagcctg cacgagcaca
     2341 ttgccaatct ggccggcagc cccgccatta agaagggcat cctgcagaca gtgaaggtgg
     2401 tggacgagct cgtgaaagtg atgggccggc acaagcccga gaacatcgtg atcgaaatgg
     2461 ccagagagaa ccagaccacc cagaagggac agaagaacag ccgcgagaga atgaagcgga
     2521 tcgaagaggg catcaaagag ctgggcagcc agatcctgaa agaacacccc gtggaaaaca
     2581 cccagctgca gaacgagaag ctgtacctgt actacctgca gaatgggcgg gatatgtacg
     2641 tggaccagga actggacatc aaccggctgt ccgactacga tgtggaccac atcgtgcctc
     2701 agagctttct gaaggacgac tccatcgaca acaaggtgct gaccagaagc gacaaggccc
     2761 ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa gaagatgaag aactactggc
     2821 ggcagctgct gaacgccaag ctgattaccc agagaaagtt cgacaatctg accaaggccg
     2881 agagaggcgg cctgagcgaa ctggataagg ccggcttcat caagagacag ctggtggaaa
     2941 cccggcagat cacaaagcac gtggcacaga tcctggactc ccggatgaac actaagtacg
     3001 acgagaatga caagctgatc cgggaagtga aagtgatcac cctgaagtcc aagctggtgt
     3061 ccgatttccg gaaggatttc cagttttaca aagtgcgcga gatcaacaac taccaccacg
     3121 cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc
     3181 tggaaagcga gttcgtgtac ggcgactaca aggtgtacga cgtgcggaag atgatcgcca
     3241 agagcgagca ggaaatcggc aaggctaccg ccaagtactt cttctacagc aacatcatga
     3301 actttttcaa gaccgagatt accctggcca acggcgagat ccggaagcgg cctctgatcg
     3361 agacaaacgg cgaaaccggg gagatcgtgt gggataaggg ccgggatttt gccaccgtgc
     3421 ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg
     3481 gcttcagcaa agagtctatc ctgcccaaga ggaacagcga taagctgatc gccagaaaga
     3541 aggactggga ccctaagaag tacggcggct tcgacagccc caccgtggcc tattctgtgc
     3601 tggtggtggc caaagtggaa aagggcaagt ccaagaaact gaagagtgtg aaagagctgc
     3661 tggggatcac catcatggaa agaagcagct tcgagaagaa tcccatcgac tttctggaag
     3721 ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa gctgcctaag tactccctgt
     3781 tcgagctgga aaacggccgg aagagaatgc tggcctctgc cggcgaactg cagaagggaa
     3841 acgaactggc cctgccctcc aaatatgtga acttcctgta cctggccagc cactatgaga
     3901 agctgaaggg ctcccccgag gataatgagc agaaacagct gtttgtggaa cagcacaagc
     3961 actacctgga cgagatcatc gagcagatca gcgagttctc caagagagtg atcctggccg
     4021 acgctaatct ggacaaagtg ctgtccgcct acaacaagca ccgggataag cccatcagag
     4081 agcaggccga gaatatcatc cacctgttta ccctgaccaa tctgggagcc cctgccgcct
     4141 tcaagtactt tgacaccacc atcgaccgga agaggtacac cagcaccaaa gaggtgctgg
     4201 acgccaccct gatccaccag agcatcaccg gcctgtacga gacacggatc gacctgtctc
     4261 agctgggagg cgacagcgct ggaggaggtg gaagcggagg aggaggaagc ggaggaggag
     4321 gtagcggacc taagaaaaag aggaaggtgg cggccgctgg atccggacgg gctgacgcat
     4381 tggacgattt tgatctggat atgctgggaa gtgacgccct cgatgatttt gaccttgaca
     4441 tgcttggttc ggatgccctt gatgactttg acctcgacat gctcggcagt gacgcccttg
     4501 atgatttcga cctggacatg ctgattaact gtacaggcag tggagagggc agaggaagtc
     4561 tgctaacatg cggtgacgtc gaggagaatc ctggcccaat ggccaagcct ttgtctcaag
     4621 aagaatccac cctcattgaa agagcaacgg ctacaatcaa cagcatcccc atctctgaag
     4681 actacagcgt cgccagcgca gctctctcta gcgacggccg catcttcact ggtgtcaatg
     4741 tatatcattt tactggggga ccttgtgcag aactcgtggt gctgggcact gctgctgctg
     4801 cggcagctgg caacctgact tgtatcgtcg cgatcggaaa tgagaacagg ggcatcttga
     4861 gcccctgcgg acggtgccga caggtgcttc tcgatctgca tcctgggatc aaagccatag
     4921 tgaaggacag tgatggacag ccgacggcag ttgggattcg tgaattgctg ccctctggtt
     4981 atgtgtggga gggctaaaaa tgtcgacggt accgcgggcc cgggatcctc tagatcataa
     5041 tcagccatac cacatttgta gaggttttac ttgctttaaa aaacctccca cacctccccc
     5101 tgaacctgaa acataaaatg aatgcaattg ttgttgttaa cttgtttatt gcagcttata
     5161 atggttacaa ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc
     5221 attctagttg tggtttgtcc aaactcatca atgtatctta atttatcaac aagtttgtac
     5281 aaaaaagctg aacgagaaac gtaaaatgat ataaatatca atatattaaa ttagattttg
     5341 cataaaaaac agactacata atactgtaaa acacaacata tccagtcact atggcggccg
     5401 cattaggcac cccaggcttt acactttatg cttccggctc gtataatgtg tggattttga
     5461 gttaggatcc gtcgagattt tcaggagcta aggaagctaa aatggagaaa aaaatcactg
     5521 gatataccac cgttgatata tcccaatggc atcgtaaaga acattttgag gcatttcagt
     5581 cagttgctca atgtacctat aaccagaccg ttcagctgga tattacggcc tttttaaaga
     5641 ccgtaaagaa aaataagcac aagttttatc cggcctttat tcacattctt gcccgcctga
     5701 tgaatgctca tccggaattc cgtatggcaa tgaaagacgg tgagctggtg atatgggata
     5761 gtgttcaccc ttgttacacc gttttccatg agcaaactga aacgttttca tcgctctgga
     5821 gtgaatacca cgacgatttc cggcagtttc tacacatata ttcgcaagat gtggcgtgtt
     5881 acggtgaaaa cctggcctat ttccctaaag ggtttattga gaatatgttt ttcgtctcag
     5941 ccaatccctg ggtgagtttc accagttttg atttaaacgt ggccaatatg gacaacttct
     6001 tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc
     6061 tggcgattca ggttcatcat gccgtttgtg atggcttcca tgtcggcaga atgcttaatg
     6121 aattacaaca gtactgcgat gagtggcagg gcggggcgta aagatctgga tccggcttac
     6181 taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata agaatatata
     6241 ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc gtattacagt
     6301 gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa tatctccggt
     6361 ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg
     6421 gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct
     6481 gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa gagagagccg
     6541 ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt
     6601 gatccccctg gccagtgcac gtctgctgtc agataaagtc tcccgtgaac tttacccggt
     6661 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt
     6721 ctccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc
     6781 cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtctgc
     6841 aggtcgacca tagtgactgg atatgttgtg ttttacagta ttatgtagtc tgttttttat
     6901 gcaaaatcta atttaatata ttgatattta tatcatttta cgtttctcgt tcagctttct
     6961 tgtacaaagt ggttgataaa tccatcgaag cagctccagt ctacacaatc gctcaagacg
     7021 tgtaatgctt ttattatata ttagtcacga tatctataac aagaaaatat atatataata
     7081 agttatcacg taagtagaac atgaaataac aatataatta tcgtatgagt taaatcttaa
     7141 aagtcacgta aaagataatc atgcgtcatt ttgactcacg cggtcgttat agttcaaaat
     7201 cagtgacact taccgcattg acaagcacgc ctcacgggag ctccaagcgg cgactgagat
     7261 gtcctaaatg cacagcgacg gattcgcgct atttagaaag agagagcaat atttcaagaa
     7321 tgcatgcgtc aattttacgc agactatctt tctagggtta aaaaagattt gcgctttact
     7381 cgacctaaac tttaaacacg tcatagaatc ttcgtttgac aaaaaccaca ttgtggggta
     7441 ccgagctcga attcatcgat gatatcagat ctgccggtct ccctatagtg agtcgtatta
     7501 atttcgataa gccaggttaa cctgcattaa tgaatcggcc aacgcgcggg gagaggcggt
     7561 ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg
     7621 ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg
     7681 gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag
     7741 gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga
     7801 cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct
     7861 ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc
     7921 tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg
     7981 gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc
     8041 tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca
     8101 ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag
     8161 ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct
     8221 ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc
     8281 accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga
     8341 tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca
     8401 cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat
     8461 taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac
     8521 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
     8581 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
     8641 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
     8701 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
     8761 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
     8821 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
     8881 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
     8941 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
     9001 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
     9061 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
     9121 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
     9181 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
     9241 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
     9301 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
     9361 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
     9421 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
     9481 cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta
     9541 acctataaaa ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt
     9601 gaaaacctct gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc
     9661 gggagcagac aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt
     9721 aactatgcgg catcagagca gattgtactg agagtgcacc atatggacat attgtcgtta
     9781 gaacgcggct acaattaata cataacctta tgtatcatac acatacgatt taggtgacac
     9841 tatagaactc gacctcgagg ctggcacgac aggtttcccg actggaaagc gggcagtgag
     9901 cgcaacgcaa ttaatgtgag ttagctcact cattaggcac cccaggcttt acactttatg
     9961 cttccggctc gtatgttgtg tggaattgtg agcggataac aatttcacac aggaaacagc
    10021 tatgaccatg attacgccaa gctcgaaatt aaccctcact aaagggaaca aaagctggag
    10081 ctcgtctttg atcaaaacgc aaatcgacga aaatgtgtcg gacaatatca agtcgatgag
    10141 cgaaaaacta aaaaggctag aatacgacaa tctcacagac agcgttgaga tatacggtat
    10201 tcacgacagc aggctgaata ataaaaaaat tagaaactat tatttaaccc tagaaagata
    10261 atcatattgt gacgtacgtt aaagataatc atgcgtaaaa ttgacgcatg tgttttatcg
    10321 atctgtatat cgaggtttat ttattaattt gaatagatat taagttttat tatatttaca
    10381 cttacatact aataataaat tcaacaaaca atttatttat gtttatttat ttattaaaaa
    10441 aaaacaaaaa ctcaaaattt cttctataaa gtaacaaaac ttttaaacat tctctctttt
    10501 acaaaaataa acttattttg tactttaaaa acagtcatgt tgtattataa aataagtaat
    10561 tagcttaact tatacataat agaaacaaat tatacttatt aattaatcgt gaggctccgg
    10621 tgcccgtcag tgggcagagc gcacatcgcc cacagtcccc gagaagttgg ggggaggggt
    10681 cggcaattga accggtgcct agagaaggtg gcgcggggta aactgggaaa gtgatgtcgt
    10741 gtactggctc cgcctttttc ccgagggtgg gggagaaccg tatataagtg cagtagtcgc
    10801 cgtgaacgtt ctttttcgca acgggtttgc cgccagaaca caggtaagtg ccgtgtgtgg
    10861 ttcccgcggg cctggcctct ttacgggtta tggcccttgc gtgccttgaa ttacttccac
    10921 ctggctgcag tacgtgattc ttgatcccga gcttcgggtt ggaagtgggt gggagagttc
    10981 gaggccttgc gcttaaggag ccccttcgcc tcgtgcttga gttgaggcct ggcctgggcg
    11041 ctggggccgc cgcgtgcgaa tctggtggca ccttcgcgcc tgtctcgctg ctttcgataa
    11101 gtctctagcc atttaaaatt tttgatgacc tgctgcgacg ctttttttct ggcaagatag
    11161 tcttgtaaat gcgggccaag atctgcacac tggtatttcg gtttttgggg ccgcgggcgg
    11221 cgacggggcc cgtgcgtccc agcgcacatg ttcggcgagg cggggcctgc gagcgcggcc
    11281 accgagaatc ggacgggggt agtctcaagc tggccggcct gctctggtgc ctggcctcgc
    11341 gccgccgtgt atcgccccgc cctgggcggc aaggctggcc cggtcggcac cagttgcgtg
    11401 agcggaaaga tggccgcttc ccggccctgc tgcagggagc tcaaaatgga ggacgcggcg
    11461 ctcgggagag cgggcgggtg agtcacccac acaaaggaaa agggcctttc cgtcctcagc
    11521 cgtcgcttca tgtgactcca cggagtaccg ggcgccgtcc aggcacctcg attagttctc
    11581 gagcttttgg agtacgtcgt ctttaggttg gggggagggg ttttatgcga tggagtttcc
    11641 ccacactgag tgggtggaga ctgaagttag gccagcttgg cacttgatgt aattctcctt
    11701 ggaatttgcc ctttttgagt ttggatcttg gttcattctc aagcctcaga cagtggttca
    11761 aagttttttt cttccatttc aggtgtcgtg aaggtttacc atggcttcaa actttactca
    11821 gttcgtgctc gtggacaatg gtgggacagg ggatgtgaca gtggctcctt ctaatttcgc
    11881 taatggggtg gcagagtgga tcagctccaa ctcacggagc caggcctaca aggtgacatg
    11941 cagcgtcagg cagtctagtg cccagaagag aaagtatacc atcaaggtgg aggtccccaa
    12001 agtggctacc cagacagtgg gcggagtcga actgcctgtc gccgcttgga ggtcctacct
    12061 gaacatggag ctcactatcc caattttcgc taccaattct gactgtgaac tcatcgtgaa
    12121 ggcaatgcag gggctcctca aagacggtaa tcctatccct tccgccatcg ccgctaactc
    12181 aggtatctac agcgctggag gaggtggaag cggaggagga ggaagcggag gaggaggtag
    12241 cggacctaag aaaaagagga aggtggcggc cgctggatcc ccttcagggc agatcagcaa
    12301 ccaggccctg gctctggccc ctagctccgc tccagtgctg gcccagacta tggtgccctc
    12361 tagtgctatg gtgcctctgg cccagccacc tgctccagcc cctgtgctga ccccaggacc
    12421 accccagtca ctgagcgctc cagtgcccaa gtctacacag gccggcgagg ggactctgag
    12481 tgaagctctg ctgcacctgc agttcgacgc tgatgaggac ctgggagctc tgctggggaa
    12541 cagcaccgat cccggagtgt tcacagatct ggcctccgtg gacaactctg agtttcagca
    12601 gctgctgaat cagggcgtgt ccatgtctca tagtacagcc gaaccaatgc tgatggagta
    12661 ccccgaagcc attacccggc tggtgaccgg cagccagcgg ccccccgacc ccgctccaac
    12721 tcccctggga accagcggcc tgcctaatgg gctgtccgga gatgaagact tctcaagcat
    12781 cgctgatatg gactttagtg ccctgctgtc acagatttcc tctagtgggc agggaggagg
    12841 tggaagcggc ttcagcgtgg acaccagtgc cctgctggac ctgttcagcc cctcggtgac
    12901 cgtgcccgac atgagcctgc ctgaccttga cagcagcctg gccagtatcc aagagctcct
    12961 gtctccccag gagcccccca ggcctcccga ggcagagaac agcagcccgg attcagggaa
    13021 gcagctggtg cactacacag cgcagccgct gttcctgctg gaccccggct ccgtggacac
    13081 cgggagcaac gacctgccgg tgctgtttga gctgggagag ggctcctact tctccgaagg
    13141 ggacggcttc gccgaggacc ccaccatctc cctgctgaca ggctcggagc ctcccaaagc
    13201 caaggacccc actgtctcct gtacaggcag tggagagggc agaggaagtc tgctaacatg
    13261 cggtgacgtc gaggagaatc ctggcccaac catgaaaaag cctgaactca ccgctacctc
    13321 tgtcgagaag tttctgatcg aaaagttcga cagcgtctcc gacctgatgc agctctccga
    13381 gggcgaagaa tctcgggctt tcagcttcga tgtgggaggg cgtggatatg tcctgcgggt
    13441 gaatagctgc gccgatggtt tctacaaaga tcgctatgtt tatcggcact ttgcatccgc
    13501 cgctctccct attcccgaag tgcttgacat tggggagttc agcgagagcc tgacctattg
    13561 catctcccgc cgtgcacagg gtgtcacctt gcaagacctg cctgaaaccg aactgcccgc
    13621 tgttctccag cccgtcgccg aggccatgga tgccatcgct gccgccgatc ttagccagac
    13681 cagcgggttc ggcccattcg gacctcaagg aatcggtcaa tacactacat ggcgcgattt
    13741 catctgcgct attgctgatc cccatgtgta tcactggcaa actgtgatgg acgacaccgt
    13801 cagtgcctcc gtcgcccagg ctctcgatga gctgatgctt tgggccgagg actgccccga
    13861 agtccggcac ctcgtgcacg ccgatttcgg ctccaacaat gtcctgaccg acaatggccg
    13921 cataacagcc gtcattgact ggagcgaggc catgttcggg gattcccaat acgaggtcgc
    13981 caacatcttc ttctggaggc cctggttggc ttgtatggag cagcagaccc gctacttcga
    14041 gcggaggcat cccgagcttg caggatctcc tcggctccgg gcttatatgc tccgcattgg
    14101 tcttgaccaa ctctatcaga gcttggttga cggcaatttc gatgatgcag cttgggctca
    14161 gggtcgctgc gacgcaatcg tccggtccgg agccgggact gtcgggcgta cacaaatcgc
    14221 ccgcagaagc gctgccgtct ggaccgatgg ctgtgtggaa gtgctcgccg atagtggaaa
    14281 cagacgcccc agcactcgtc ctagggcaaa ggatctgcag taaaaacgat cataatcagc
    14341 cataccacat ttgtagaggt tttacttgct ttaaaaaacc tcccacacct ccccctgaac
    14401 ctgaaacata aaatgaatgc aattgtcgtt gttaacttgt ttattgcagc ttataatggt
    14461 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
    14521 agttgtggtt tgtccaaact catcaatgta tctta
//
