LOCUS       sequence-428841-       15550 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gene expression by Piggybac transposase in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15550)
  TITLE     Yoshimatsu Lab plasmids 2023
REFERENCE   2  (bases 1 to 15550)
  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"
COMMENT     Sequence Label: pPB-CAG-Brainbow4.2-PGK-HygTK
FEATURES             Location/Qualifiers
     source          1..15550
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        1012..1030
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     repeat_region   1054..1116
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     CDS             complement(1642..2637)
                     /codon_start=1
                     /product="truncated herpes simplex virus thymidine kinase 
                     (Salomon et al., 1995)"
                     /label=Delta-TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWRV
                     LGASETIANIYTTQHRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGS
                     SHAPPPALTLIFDRHPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGAL
                     PEDRHIDRLAKRQRPGERLDLAMLAAIRRVYGLLANTVRYLQCGGSWREDWGQLSGTAV
                     PPQGAEPQSNAGPRPHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRSMHVFI
                     LDYDQSPAGCRDALLQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     promoter        complement(3726..4225)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(3774..3793)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     4167..4190
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     polyA_signal    4388..4595
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    4615..4639
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(4678..4711)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     polyA_signal    4724..4845
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    4979..5567
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             5085..5096
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(5655..6350)
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     regulatory      6347..6356
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    6365..6398
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     polyA_signal    6417..6538
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    6672..7260
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             6778..6789
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(join(7348..8058,8059..8061,8062..8064))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(7348..8058,8059..8061,8062..8064))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      8061..8070
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    8079..8112
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     polyA_signal    8129..8250
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    8384..8972
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             8490..8501
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(9060..9767)
                     /codon_start=1
                     /product="photostable monomeric orange derivative of DsRed 
                     fluorescent protein (Shaner et al., 2008)"
                     /label=mOrange2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEENNMAIIKEFMRFKVRMEGSVNGHEFEIEGEGEGRPYEG
                     FQTAKLKVTKGGPLPFAWDILSPHFTYGSKAYVKHPADIPDYFKLSFPEGFKWERVMNY
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GKIKMRLKLKDGGHYTSEVKTTYKAKKPVQLPGAYIVDIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     regulatory      9764..9773
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    9782..9815
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     polyA_signal    9828..9949
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    10143..10731
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             10249..10260
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     CDS             complement(join(10737..11450,11451..11453,11454..11456))
                     /codon_start=1
                     /product="enhanced monomeric variant of CFP (Rizzo et al., 
                     2004)"
                     /label=mCerulean
                     /note=" mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLSWGVQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNAIHGNVYITADKQKNGIK
                     ANFGLNCNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      11453..11462
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    11471..11504
                     /label=loxN
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AGGTATAC) (Shaw et al., 2021). loxN sites are 
                     compatible with each other, but incompatible with loxP or 
                     lox2272 sites."
     protein_bind    11513..11546
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    11555..11588
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(11637..11661)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(11710..11729)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     intron          11737..12754
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     complement(11791..11813)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     promoter        complement(12755..13032)
                     /label=chicken beta-actin promoter
     enhancer        13037..13420
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     repeat_region   13698..13732
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(13750..13769)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(13751..13769)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(13776..13793)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(13776..13792)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(13785..13807)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      13934..14389
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(14021..14040)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     14231..14252
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        14415..14519
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(14520..14588,14589..15380)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(14738..14757)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctcg aaattaaccc
     1021 tcactaaagg gaacaaaagc tggagctctt taaccctaga aagataatca tattgtgacg
     1081 tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
     1141 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1201 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca
     1261 aaatttcttc tataaagtaa caaaggatct gatggatcta tagatcatga gtgggaggaa
     1321 tgagctggcc cttaatttgg ttttgcttgt ttaaattatg atatccaact atgaaacatt
     1381 atcataaagc aatagtaaag agccttcagt aaagagcagg catttatcta atcccacccc
     1441 acccccaccc ccgtagctcc aatccttcca ttcaaaatgt aggtactctg ttctcaccct
     1501 tcttaacaaa gtatgacagg aaaaacttcc atgtggacat ctttattgtt taatagatca
     1561 tcaatttctc gactattcct ttgccctcgg acgagtgctg gggcgtcggt ttccactatc
     1621 ggcgagtcga taagctagag ctcagttagc ctcccccatc tcccgggcaa acgtgcgcgc
     1681 caggtcgcat atcgtcggta tggagccggg ggtggtgacg tgggtctgga ccatcccgga
     1741 ggtaagttgc agcagggcgt cccggcagcc ggcgggcgat tggtcgtaat ccaggataaa
     1801 gacgtgcatg gaacggaggc gtttggccaa gacgtccaag gcccaggcaa acacgttata
     1861 caggtcgccg ttgggggcca gcaactcggg ggcccgaaac agggtaaata acgtgtcccc
     1921 gatatggggt cgtgggcccg cgttgctctg gggctcggca ccctggggcg gcacggccgt
     1981 ccccgaaagc tgtccccagt cctcccgcca cgacccgccg cactgcagat accgcaccgt
     2041 attggcaagt agcccgtaaa cgcggcgaat cgcagccagc atagccaggt ccagccgctc
     2101 gccggggcgc tggcgtttgg ccaggcggtc gatgtgtctg tcctccggaa gggccccaag
     2161 cacgatgttg gtgccgggca aggtcggcgg gatgagggcc acgaacgcca gcacggcctg
     2221 gggggtcatg ctgcccataa ggtaccgcgc ggccgggtag cacaggaggg cggcgatggg
     2281 atggcggtcg aagatgaggg tgagggccgg gggcggggca tgtgagctcc cagcctcccc
     2341 cccgatatga ggagccagaa cggcgtcggt cacggcataa ggcatgccca ttgttatctg
     2401 ggcgcttgtc attaccaccg ccgcgtcccc ggccgatatc tcaccctggt cgaggcggtg
     2461 ttgtgtggtg tagatgttcg cgattgtctc ggaagccccc agcacccgcc agtaagtcat
     2521 cggctcgggt acgtagacga tatcgtcgcg cgaacccagg gccaccagca gttgcgtggt
     2581 ggtggttttc cccatcccgt ggggaccgtc tatataaacc cgcagtagcg tgggcatttt
     2641 ctgctccggg cggacttccg tggcttcttg ctgccggcga gggcgcaacg ccgtacgtcg
     2701 gttgctatgg ccgcgagaac gcgcagcctg gtcgaacgca gacgcgactt ctacacagcc
     2761 atcggtccag acggccgcgc ttctgcgggc gatttgtgta cgcccgacag tcccggctcc
     2821 ggatcggacg attgcgtcgc atcgaccctg cgcccaagct gcatcatcga aattgccgtc
     2881 aaccaagctc tgatagagtt ggtcaagacc aatgcggagc atatacgccc ggagccgcgg
     2941 cgatcctgca agctccggat gcctccgctc gaagtagcgc gtctgctgct ccatacaagc
     3001 caaccacggc ctccagaaga agatgttggc gacctcgtat tgggaatccc cgaacatcgc
     3061 ctcgctccag tcaatgaccg ctgttatgcg gccattgtcc gtcaggacat tgttggagcc
     3121 gaaatccgcg tgcacgaggt gccggacttc ggggcagtcc tcggcccaaa gcatcagctc
     3181 atcgagagcc tgcgcgacgg acgcactgac ggtgtcgtcc atcacagttt gccagtgata
     3241 cacatgggga tcagcaatcg cgcatatgaa atcacgccat gtagtgtatt gaccgattcc
     3301 ttgcggtccg aatgggccga acccgctcgt ctggctaaga tcggccgcag cgatcgcatc
     3361 catggcctcc gcgaccggct gcagaacagc gggcagttcg gtttcaggca ggtcttgcaa
     3421 cgtgacaccc tgtgcacggc gggagatgca ataggtcagg ctctcgctga attccccaat
     3481 gtcaagcact tccggaatcg ggagcgcggc cgatgcaaag tgccgataaa cataacgatc
     3541 tttgtagaaa ccatcggcgc agctatttac ccgcaggaca tatccacgcc ctcctacatc
     3601 gaagctgaaa gcacgagatt cttcgccctc cgagagctgc atcaggtcgg agacgctgtc
     3661 gaacttttcg atcagaaact tctcgacaga cgtcgcggtg agttcaggct ttttcatgag
     3721 gatcacgaaa ggcccggaga tgaggaagag gagaacagcg cggcagacgt gcgcttttga
     3781 agcgtgcaga atgccgggcc tccggaggac cttcgggcgc ccgccccgcc cctgagcccg
     3841 cccctgagcc cgcccccgga cccacccctt cccagcctct gagcccagaa agcgaaggag
     3901 caaagctgct attggccgct gccccaaagg cctacccgct tccattgctc agcggtgctg
     3961 tccatctgca cgagactagt gagacgtgct acttccattt gtcacgtcct gcacgacgcg
     4021 agctgcgggg cgggggggaa cttcctgact aggggaggag tagaaggtgg cgcgaagggg
     4081 ccaccaaaga acggagccgg ttggcgccta ccggtggatg tggaatgtgt gcgaggccag
     4141 aggccacttg tgtagcgcca agtgcccagc ggggctgcta aagcgcatgc tccagactgc
     4201 cttgggaaaa gcgcctcccc tacccggtag aattccccgc aagaggcccg gcagtaccgg
     4261 cataaccaag cctatgccta cagcatccag ggtgacggtg ccgaggatga cgatgagcgc
     4321 attgttagat ttcatacacg gtgcctgact gcgttagcaa tttaactgtg ataaactacc
     4381 gcattaatcc ccagcatgcc tgctattctc ttcccaatcc tcccccttgc tgtcctgccc
     4441 caccccaccc cccagaatag aatgacacct actcagacaa tgcgatgcaa tttcctcatt
     4501 ttattaggaa aggacagtgg gagtggcacc ttccagggtc aaggaaggca cgggggaggg
     4561 gcaaacaaca gatggctggc aactagaagg cacagtgcat gcctgcaggt cggggccact
     4621 ttgtacaaga aagctgggtc tagatgcatt cgctacctta ggaccgttat agttacggaa
     4681 gttcctatac tttctagaga ataggaactt cactagtacg ccttaagata cattgatgag
     4741 tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga aatttgtgat
     4801 gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa caacaattgc
     4861 attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag caagtaaaac
     4921 ctctacaaat gtggtatggc tgattatgat ctggtgcagc agtctcgacg gtatcgatgc
     4981 ggggaggcgg cccaaaggga gatccgactc gtctgagggc gaaggcgaag acgcggaaga
     5041 ggccgcagag ccggcagcag gccgcgggaa ggaaggtccg ctggattgag ggccgaaggg
     5101 acgtagcaga aggacgtccc gcgcagaatc caggtggcaa cacaggcgag cagccaagga
     5161 aaggacgatg atttccccga caacaccacg gaattgtcag tgcccaacag ccgagcccct
     5221 gtccagcagc gggcaaggca ggcggcgatg agttccgccg tggcaatagg gagggggaaa
     5281 gcgaaagtcc cggaaaggag ctgacaggtg gtggcaatgc cccaaccagt gggggttgcg
     5341 tcagcaaaca cagtgcacac cacgccacgt tgcctgacaa cgggccacaa ctcctcataa
     5401 agagacagca accaggattt atacaaggag gagaaaatga aagccatacg ggaagcaata
     5461 gcatgataca aaggcattaa agcagcgtat ccacatagcg taaaaggagc aacatagtta
     5521 agaataccag tcaatctttc acaaattttg taatccagag gttgattagc ggccgctcag
     5581 gagagcacac acttgcagct catgcagccg gggccactct catcaggagg gttcagctta
     5641 ctcacgcgtg cacttctgtg ccccagtttg ctagggaggt cgcagtatct ggccacagcc
     5701 acctcgtgct gctcgacgta ggtctctttg tcggcctcct tgattctttc cagtcttctg
     5761 tccacatagt agacgccggg catcttgagg ttcttagcgg gtttcttgga tctgtatgtg
     5821 gtcttcaagt tgcagatcag gtggcccccg cccacgagct tcagggccat gtcggctctg
     5881 ccttccaggc cgccgtcagc ggggtacagg gtctcggtgg aggcctccca gccgagtgtt
     5941 ttcttctgca tcacagggcc gttggatggg aagttcaccc ctctgatctt gacgttgtag
     6001 atgaggcagc cgtcctggag gctggtgtcc tgggtagcgg tcagcacgcc cccgtcttcg
     6061 tatgtggtga ctctctccca tgtgaagccc tcggggaagg actgcttaaa gaagtcgggg
     6121 atgccctggg tgtggttgat gaaggttttg ctgccgtaca tgaagctggt agccaggatg
     6181 tcgaaggcga aggggagagg gccgccctcg accgccttga ttctcatggt ctgggtgccc
     6241 tcgtagggct tgccttcgcc ctcggatgtg cacttgaagt ggtggttgtt cacggtgccc
     6301 tccatgtaca gcttcatgtg catgttctcc ttaatcagct cgctcaccat ggtggcagta
     6361 cttgataact tcgtataagg tatactatac gaagttatat gaagatcttc acgccttaag
     6421 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     6481 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     6541 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta
     6601 aagcaagtaa aacctctaca aatgtggtat ggctgattat gatctggtgc agcagtctcg
     6661 acggtatcga tgcggggagg cggcccaaag ggagatccga ctcgtctgag ggcgaaggcg
     6721 aagacgcgga agaggccgca gagccggcag caggccgcgg gaaggaaggt ccgctggatt
     6781 gagggccgaa gggacgtagc agaaggacgt cccgcgcaga atccaggtgg caacacaggc
     6841 gagcagccaa ggaaaggacg atgatttccc cgacaacacc acggaattgt cagtgcccaa
     6901 cagccgagcc cctgtccagc agcgggcaag gcaggcggcg atgagttccg ccgtggcaat
     6961 agggaggggg aaagcgaaag tcccggaaag gagctgacag gtggtggcaa tgccccaacc
     7021 agtgggggtt gcgtcagcaa acacagtgca caccacgcca cgttgcctga caacgggcca
     7081 caactcctca taaagagaca gcaaccagga tttatacaag gaggagaaaa tgaaagccat
     7141 acgggaagca atagcatgat acaaaggcat taaagcagcg tatccacata gcgtaaaagg
     7201 agcaacatag ttaagaatac cagtcaatct ttcacaaatt ttgtaatcca gaggttgatt
     7261 agcggccgct caggagagca cacacttgca gctcatgcag ccggggccac tctcatcagg
     7321 agggttcagc ttactcacgc gtgcactctt gtacagctcg tccatgccga gagtgatccc
     7381 ggcggcggtc acgaactcca gcaggaccat gtgatcgcgc ttctcgttgg ggtctttgct
     7441 cagggcggac tgggtgctca ggtagtggtt gtcgggcagc agcacggggc cgtcgccgat
     7501 gggggtgttc tgctggtagt ggtcggcgag ctgcacgctg ccgtcctcga tgttgtggcg
     7561 gatcttgaag ttcaccttga tgccgttctt ctgcttgtcg gccatgatat agacgttgtg
     7621 gctgttgtag ttgtactcca gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat
     7681 gcccttcagc tcgatgcggt tcaccagggt gtcgccctcg aacttcacct cggcgcgggt
     7741 cttgtagttg ccgtcgtcct tgaagaagat ggtgcgctcc tggacgtagc cttcgggcat
     7801 ggcggacttg aagaagtcgt gctgcttcat gtggtcgggg tagcggctga agcactgcac
     7861 gccgtaggtc agggtggtca cgagggtggg ccagggcacg ggcagcttgc cggtggtgca
     7921 gatgaacttc agggtcagct tgccgtaggt ggcatcgccc tcgccctcgc cggacacgct
     7981 gaacttgtgg ccgtttacgt cgccgtccag ctcgaccagg atgggcacca ccccggtgaa
     8041 cagctcctcg cccttgctca ccatggtggc cgtacgcgat aacttcgtat aaagtatcct
     8101 atacgaagtt atatgtttaa acacgcctta agatacattg atgagtttgg acaaaccaca
     8161 actagaatgc agtgaaaaaa atgctttatt tgtgaaattt gtgatgctat tgctttattt
     8221 gtaaccatta taagctgcaa taaacaagtt aacaacaaca attgcattca ttttatgttt
     8281 caggttcagg gggaggtgtg ggaggttttt taaagcaagt aaaacctcta caaatgtggt
     8341 atggctgatt atgatctggt gcagcagtct cgacggtatc gatgcgggga ggcggcccaa
     8401 agggagatcc gactcgtctg agggcgaagg cgaagacgcg gaagaggccg cagagccggc
     8461 agcaggccgc gggaaggaag gtccgctgga ttgagggccg aagggacgta gcagaaggac
     8521 gtcccgcgca gaatccaggt ggcaacacag gcgagcagcc aaggaaagga cgatgatttc
     8581 cccgacaaca ccacggaatt gtcagtgccc aacagccgag cccctgtcca gcagcgggca
     8641 aggcaggcgg cgatgagttc cgccgtggca atagggaggg ggaaagcgaa agtcccggaa
     8701 aggagctgac aggtggtggc aatgccccaa ccagtggggg ttgcgtcagc aaacacagtg
     8761 cacaccacgc cacgttgcct gacaacgggc cacaactcct cataaagaga cagcaaccag
     8821 gatttataca aggaggagaa aatgaaagcc atacgggaag caatagcatg atacaaaggc
     8881 attaaagcag cgtatccaca tagcgtaaaa ggagcaacat agttaagaat accagtcaat
     8941 ctttcacaaa ttttgtaatc cagaggttga ttagcggccg ctcaggagag cacacacttg
     9001 cagctcatgc agccggggcc actctcatca ggagggttca gcttactcac gcgtgcactc
     9061 ttgtacagct cgtccatgcc gccggtggag tggcggccct cggcgcgttc gtactgttcc
     9121 acgatggtgt agtcctcgtt gtgggaggtg atgtccaact tgatgtcgac gatgtaggcg
     9181 ccgggcagct gcacgggctt cttggccttg taggtggtct tgacctcgga ggtgtagtgg
     9241 ccgccgtcct tcagcttcag cctcatcttg atcttgccct tcagggcacc gtcctcgggg
     9301 tacatccgct cggaggaggc ctcccagccc atggtcttct tctgcatcac ggggccgtcg
     9361 gaggggaagt tggtgccgcg cagcttcacc ttgtagatga actcgccgtc ctgcagggag
     9421 gagtcctggg tcacggtcac cacgccgccg tcctcgtagt tcatcacgcg ctcccacttg
     9481 aagccctcgg ggaaggacag cttgaagtag tcggggatgt cggcggggtg cttcacgtag
     9541 gccttggagc cgtaggtgaa atgaggggac aggatgtccc aggcgaaggg cagggggcca
     9601 cccttggtca ccttcagctt agcggtctga aagccctcgt aggggcggcc ctcgccctcg
     9661 ccctcgatct cgaactcgtg gccgttcacg gagccctcca tgcgcacctt gaagcgcatg
     9721 aactccttga tgatggccat gttattctcc tcgcccttgc tcaccatggt ggcaacgtta
     9781 gataacttcg tataatgtat gctatacgaa gttatgctag cacgccttaa gatacattga
     9841 tgagtttgga caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg
     9901 tgatgctatt gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa
     9961 ttgcattcat tttatgtttc aggttcaggg ggaggtgtgg gaggtttttt aaagcaagta
    10021 aaacctctac aaatgtggta tggctgatta tgatctagag tcgcggccag atggagttct
    10081 gaggtcatta ctggatctat caacaggagt ccaagctcag ctaattaagc taatttaccg
    10141 atgcggggag gcggcccaaa gggagatccg actcgtctga gggcgaaggc gaagacgcgg
    10201 aagaggccgc agagccggca gcaggccgcg ggaaggaagg tccgctggat tgagggccga
    10261 agggacgtag cagaaggacg tcccgcgcag aatccaggtg gcaacacagg cgagcagcca
    10321 tggaaaggac gtcagcttcc ccgacaacac cacggaattg tcagtgccca acagccgagc
    10381 ccctgtccag cagcgggcaa ggcaggcggc gatgagttcc gccgtggcaa tagggagggg
    10441 gaaagcgaaa gtcccggaaa ggagctgaca ggtggtggca atgccccaac cagtgggggt
    10501 tgcgtcagca aacacagtgc acaccacgcc acgttgcctg acaacgggcc acaactcctc
    10561 ataaagagac agcaaccagg atttatacaa ggaggagaaa atgaaagcca tacgggaagc
    10621 aatagcatga tacaaaggca ttaaagcagc gtatccacat agcgtaaaag gagcaacata
    10681 gttaagaata ccagtcaatc tttcacaaat tttgtaatcc agaggttgat taagctttac
    10741 ttgtacagct cgtccatgcc gagagtgatc ccggcggcgg tcacgaactc cagcaggacc
    10801 atgtgatcgc gcttctcgtt ggggtctttg ctcagcttgg actgggtgct caggtagtgg
    10861 ttgtcgggca gcagcacggg gccgtcgccg atgggggtgt tctgctggta gtggtcggcg
    10921 agctgcacgc tgccgtcctc gatgttgcag ttgaggccga agttggcctt gatgccgttc
    10981 ttctgcttgt cggcggtgat atagacgttg ccgtggatgg cgttgtactc cagcttgtgc
    11041 cccaggatgt tgccgtcctc cttgaagtcg atgcccttca gctcgatgcg gttcaccagg
    11101 gtgtcgccct cgaacttcac ctcggcgcgg gtcttgtagt tgccgtcgtc cttgaagaag
    11161 atggtgcgct cctggacgta gccttcgggc atggcggact tgaagaagtc gtgctgcttc
    11221 atgtggtcgg ggtagcgggc gaagcactgc acgccccagc tcagggtggt cacgagggtg
    11281 ggccagggca cgggcagctt gccggtggtg cagatgaact tcagggtcag cttgccgtag
    11341 gtggcatcgc cctcgccctc gccggacacg ctgaacttgt ggccgtttac gtcgccgtcc
    11401 agctcgacca ggatgggcac caccccggtg aacagctcct cgcccttgct caccatggtg
    11461 gcgatatcag ataacttcgt ataaggtata ctatacgaag ttatagcgct tgataacttc
    11521 gtataaagta tcctatacga agttatgcgc gccgataact tcgtataatg tatgctatac
    11581 gaagttatcc taggttcgct accttaggac cgttatagtt acgtgcgttc ctttaaagcc
    11641 tgcttttttg tacaaacttg aattctttgc caaaatctcg aggaattctt tgccaaaatg
    11701 atgagacagc acaataacca gcacgttgcc caggagctgt aggaaaaaga agaaggcatg
    11761 aacatggtta gcagaggctc tagagccgcc ggtcacacgc cagaagccga accccgccct
    11821 gccccgtccc ccccgaaggc agccgtcccc ccgcggacag ccccgaggct ggagagggag
    11881 aaggggacgg cggcgcggcg acgcacgaag gccctccccg cccatttcct tcctgccggc
    11941 gccgcaccgc ttcgccccgc gcccgctaga gggggtgcgg cggcgcctcc cagatttcgg
    12001 ctccgcacag atttgggaca aaggaagtcc ctgcgccctc tcgcacgatt accataaaag
    12061 gcaatggctg cggctcgccg cgcctcgaca gccgccggcg ctccgggggc cgccgcgccc
    12121 ctcccccgag ccctccccgg cccgaggcgg ccccgccccg cccggcaccc ccacctgccg
    12181 ccaccccccg cccggcacgg cgagccccgc gccacgcccc gtacggagcc ccgcacccga
    12241 agccgggccg tgctcagcaa ctcggggagg ggggtgcagg gggggttgca gcccgaccga
    12301 cgcgcccaca ccccctgctc acccccccac gcacacaccc cgcacgcagc ctttgttccc
    12361 ctcgcagccc cccccgcacc gcggggcacc gcccccggcc gcgctcccct cgcgcacact
    12421 gcggagcgca caaagccccg cgccgcgccc gcagcgctca cagccgccgg gcagcgcgga
    12481 gccgcacgcg gcgctcccca cgcacacaca cacgcacgca cccccccgag ccgctccccc
    12541 cgcacaaagg gccctcccgg agcccctcaa ggctttcacg cagccacaga aaagaaacaa
    12601 gccgtcatta aaccaagcgc taattacagc ccggaggaga agggccgtcc cgcccgctca
    12661 cctgtgggag taacgcggtc agtcagagcc ggggcgggcg gcgcgaggcg gcggcggagc
    12721 ggggcacggg gcgaaggcag cgcgcagcga ctcccgcccg ccgcgcgctt cgctttttat
    12781 agggccgccg ccgccgccgc ctcgccataa aaggaaactt tcggagcgcg ccgctctgat
    12841 tggctgccgc cgcacctctc cgcctcgccc cgccccgccc ctcgccccgc cccgccccgc
    12901 ctggcgcgcg cccccccccc cccccccgcc cccatcgctg cacaaaataa ttaaaaaata
    12961 aataaataca aaattggggg tggggagggg gggagatggg gagagtgaag cagaacgtgg
    13021 ggctcacctc gaccatcatg gtaatagcga tgactaatac gtagatgtac tgccaagtag
    13081 gaaagtccca taaggtcatg tactgggcat aatgccaggc gggccattta ccgtcattga
    13141 cgtcaatagg gggcgtactt ggcatatgat acacttgatg tactgccaag tgggcagttt
    13201 accgtaaata ctccacccat tgacgtcaat ggaaagtccc tattggcgtt actatgggaa
    13261 catacgtcat tattgacgtc aatgggcggg ggtcgttggg cggtcagcca ggcgggccat
    13321 ttaccgtaag ttatgtaacg cggaactcca tatatgggct atgaactaat gaccccgtaa
    13381 ttgattacta ttaataacta gctagtcaat aatcaatgtc gacgatatct ataacaagaa
    13441 aatatatata taataagtta tcacgtaagt agaacatgaa ataacaatat aattatcgta
    13501 tgagttaaat cttaaaagtc acgtaaaaga taatcatgcg tcattttgac tcacgcggtc
    13561 gttatagttc aaaatcagtg acacttaccg cattgacaag cacgcctcac gggagctcca
    13621 agcggcgact gagatgtcct aaatgcacag cgacggattc gcgctattta gaaagagaga
    13681 gcaatatttc aagaatgcat gcgtcaattt tacgcagact atctttctag ggttaaggta
    13741 cccaattcgc cctatagtga gtcgtattac aattcactgg ccgtcgtttt acaacgtcgt
    13801 gactgggaaa accctggcgt tacccaactt aatcgccttg cagcacatcc ccctttcgcc
    13861 agctggcgta atagcgaaga ggcccgcacc gatcgccctt cccaacagtt gcgcagcctg
    13921 aatggcgaat gggacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
    13981 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
    14041 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
    14101 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
    14161 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
    14221 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
    14281 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
    14341 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt
    14401 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
    14461 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
    14521 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
    14581 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
    14641 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
    14701 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
    14761 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
    14821 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
    14881 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
    14941 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
    15001 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
    15061 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
    15121 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
    15181 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
    15241 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
    15301 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
    15361 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
    15421 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
    15481 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
    15541 aaggatcttc
//
