LOCUS       sequence-213310-       13500 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  dCas9 fused to BFP and the human DNMT1 catalytic domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13500)
  AUTHORS   Lin L, Liu Y, Xu F, Huang J, Daugaard TF, Petersen TS, Hansen B, Ye 
            L, Zhou Q, Fang F, Yang L, Li S, Floe L, Jensen KT, Shrock E, Chen 
            F, Yang H, Wang J, Liu X, Xu X, Bolund L, Nielsen AL, Luo Y
  TITLE     Genome-wide determination of on-target and off-target 
            characteristics for RNA-guided DNA methylation by dCas9 
            methyltransferases.
  JOURNAL   Gigascience. 2018 Mar 1;7(3):1-19. doi: 10.1093/gigascience/giy011.
  PUBMED    29635374
REFERENCE   2  (bases 1 to 13500)
  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"; doi: "10"; journalName: 
            "Gigascience"; date: "2018-03-1- 1"; volume: "7"; issue: "3"; pages:
            "1-19"
COMMENT     Sequence Label: pCCL-PGK-SPdCas9-BFP-DNMT1
FEATURES             Location/Qualifiers
     source          1..13500
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        82..461
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        462..665
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     616..636
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             680..860
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    907..1032
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1525..1758
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1943..1987
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    2254..2371
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2410..2909
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(2445..2468)
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     primer_bind     2842..2861
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     regulatory      2968..2977
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2974..3000,3001..3003,3004..5490,5491..5493,
                     5494..7077)
                     /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 H840A mutation in the HNH 
                     catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             2974..7077
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7081..7107
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             7087..7113
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             7126..7146
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7153..7173
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7213..7908
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     misc_feature    9445..10033
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9498..9518)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9916..9927)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             10122..10355
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     promoter        complement(10384..10402)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(10385..10402)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(10648..10667)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(10649..10667)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(10677..10694)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(10677..10693)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10686..10708)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      10835..11290
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10922..10941)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11132..11153
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        11316..11420
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(11421..11489,11490..12281)
                     /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(11639..11658)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      12452..13040
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     12941..12960
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     13194..13211
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    13328..13349
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(13364..13369,13370..13387,13388..13394)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    13402..13418
                     /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     13407..13429
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     13426..13442
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     13426..13442
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     13461..13481
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        13463..13481
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
ORIGIN
        1 aagcttggcc attgcatacg ttgtatccat atcataatat gtacatttat attggctcat
       61 gtccaacatt accgccatgt tgacattgat tattgactag ttattaatag taatcaatta
      121 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      181 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      241 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      301 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      361 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      421 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      481 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      541 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      601 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      661 gagctcgttt agtgaaccgg ggtctctctg gttagaccag atctgagcct gggagctctc
      721 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt
      781 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc
      841 agtgtggaaa atctctagca gtggcgcccg aacagggact tgaaagcgaa agggaaacca
      901 gaggagctct ctcgacgcag gactcggctt gctgaagcgc gcacggcaag aggcgagggg
      961 cggcgactgg tgagtacgcc aaaaattttg actagcggag gctagaagga gagagatggg
     1021 tgcgagagcg tcagtattaa gcgggggaga attagatcgc gatgggaaaa aattcggtta
     1081 aggccagggg gaaagaaaaa atataaatta aaacatatag tatgggcaag cagggagcta
     1141 gaacgattcg cagttaatcc tggcctgtta gaaacatcag aaggctgtag acaaatactg
     1201 ggacagctac aaccatccct tcagacagga tcagaagaac ttagatcatt atataataca
     1261 gtagcaaccc tctattgtgt gcatcaaagg atagagataa aagacaccaa ggaagcttta
     1321 gacaagatag aggaagagca aaacaaaagt aagaccaccg cacagcaagc ggccgctgat
     1381 cttcagacct ggaggaggag atatgaggga caattggaga agtgaattat ataaatataa
     1441 agtagtaaaa attgaaccat taggagtagc acccaccaag gcaaagagaa gagtggtgca
     1501 gagagaaaaa agagcagtgg gaataggagc tttgttcctt gggttcttgg gagcagcagg
     1561 aagcactatg ggcgcagcgt caatgacgct gacggtacag gccagacaat tattgtctgg
     1621 tatagtgcag cagcagaaca atttgctgag ggctattgag gcgcaacagc atctgttgca
     1681 actcacagtc tggggcatca agcagctcca ggcaagaatc ctggctgtgg aaagatacct
     1741 aaaggatcaa cagctcctgg ggatttgggg ttgctctgga aaactcattt gcaccactgc
     1801 tgtgccttgg aatgctagtt ggagtaataa atctctggaa cagatttgga atcacacgac
     1861 ctggatggag tgggacagag aaattaacaa ttacacaagc ttaatacact ccttaattga
     1921 agaatcgcaa aaccagcaag aaaagaatga acaagaatta ttggaattag ataaatgggc
     1981 aagtttgtgg aattggttta acataacaaa ttggctgtgg tatataaaat tattcataat
     2041 gatagtagga ggcttggtag gtttaagaat agtttttgct gtactttcta tagtgaatag
     2101 agttaggcag ggatattcac cattatcgtt tcagacccac ctcccaaccc cgaggggacc
     2161 cgacaggccc gaaggaatag aagaagaagg tggagagaga gacagagaca gatccattcg
     2221 attagtgaac ggatctcgac ggtatcggtt aacttttaaa agaaaagggg ggattggggg
     2281 gtacagtgca ggggaaagaa tagtagacat aatagcaaca gacatacaaa ctaaagaatt
     2341 acaaaaacaa attacaaaaa ttcaaaattt tatcgatcac gagactagcc tcgacgatgg
     2401 tcgagtaccg ggtaggggag gcgcttttcc caaggcagtc tggagcatgc gctttagcag
     2461 ccccgctggg cacttggcgc tacacaagtg gcctctggcc tcgcacacat tccacatcca
     2521 ccggtaggcg ccaaccggct ccgttctttg gtggcccctt cgcgccacct tctactcctc
     2581 ccctagtcag gaagttcccc cccgccccgc agctcgcgtc gtgcaggacg tgacaaatgg
     2641 aagtagcacg tctcactagt ctcgtgcaga tggacagcac cgctgagcaa tggaagcggg
     2701 taggcctttg gggcagcggc caatagcagc tttgctcctt cgctttctgg gctcagaggc
     2761 tgggaagggg tgggtccggg ggcgggctca ggggcgggct caggggcggg gcgggcgccc
     2821 gaaggtcctc cggaggcccg gcattctgca cgcttcaaaa gcgcacgtct gccgcgctgt
     2881 tctcctcttc ctcatctccg ggcctttcga cctctagcgg gatccggcgc gtctgcagga
     2941 tatcaagctt gcggtaccgc gggcccggcc accatggaca agaagtacag catcggcctg
     3001 gccatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc
     3061 aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa cctgatcggc
     3121 gccctgctgt tcgacagcgg agaaacagcc gaggccaccc ggctgaagag aaccgccaga
     3181 agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt cagcaacgag
     3241 atggccaagg tggacgacag cttcttccac agactggaag agtccttcct ggtggaagag
     3301 gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt ggcctaccac
     3361 gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac cgacaaggcc
     3421 gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg ccacttcctg
     3481 atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat ccagctggtg
     3541 cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag
     3601 gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat cgcccagctg
     3661 cccggcgaga agaagaatgg cctgttcggc aacctgattg ccctgagcct gggcctgacc
     3721 cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct gagcaaggac
     3781 acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta cgccgacctg
     3841 tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct gagagtgaac
     3901 accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga cgagcaccac
     3961 caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa gtacaaagag
     4021 attttcttcg accagagcaa gaacggctac gccggctaca tcgatggcgg agccagccag
     4081 gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac cgaggaactg
     4141 ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga caacggcagc
     4201 atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca ggaagatttt
     4261 tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc
     4321 tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac cagaaagagc
     4381 gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc cagcgcccag
     4441 agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa ggtgctgccc
     4501 aagcacagcc tgctgtacga gtacttcacc gtgtacaacg agctgaccaa agtgaaatac
     4561 gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa agccatcgtg
     4621 gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga ggactacttc
     4681 aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg gttcaacgcc
     4741 tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt cctggacaat
     4801 gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt tgaggacaga
     4861 gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa agtgatgaag
     4921 cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct gatcaacggc
     4981 atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga cggcttcgcc
     5041 aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga ggacatccag
     5101 aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa tctggccggc
     5161 agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga gctcgtgaaa
     5221 gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga gaaccagacc
     5281 acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa
     5341 gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct gcagaacgag
     5401 aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca ggaactggac
     5461 atcaaccggc tgtccgacta cgatgtggac gctatcgtgc ctcagagctt tctgaaggac
     5521 gactccatcg ataacaaagt gctgactcgg agcgacaaga accggggcaa gagcgacaac
     5581 gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcgccagct gctgaatgcc
     5641 aagctgatta cccagaggaa gttcgacaat ctgaccaagg ccgagagagg cggcctgagc
     5701 gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca gatcacaaag
     5761 cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa cgacaaactg
     5821 atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt ccggaaggat
     5881 ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga cgcctacctg
     5941 aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag cgagttcgtg
     6001 tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc
     6061 ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt caagaccgag
     6121 attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa cggcgaaaca
     6181 ggcgagatcg tgtgggataa gggccgggac tttgccaccg tgcggaaagt gctgtctatg
     6241 ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag caaagagtct
     6301 atcctgccca agaggaacag cgacaagctg atcgccagaa agaaggactg ggaccctaag
     6361 aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt ggccaaagtg
     6421 gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat caccatcatg
     6481 gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg ctacaaagaa
     6541 gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct ggaaaacggc
     6601 cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc
     6661 tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa gggctccccc
     6721 gaggataatg agcagaaaca gctgtttgtg gaacagcaca aacactacct ggacgagatc
     6781 atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa tctggacaag
     6841 gtgctgagcg cctacaacaa gcacagagac aagcctatca gagagcaggc cgagaatatc
     6901 atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta ctttgacacc
     6961 accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac cctgatccac
     7021 cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg aggcgacgcc
     7081 tatccctatg acgtgcccga ttatgccagc ctgggcagcg gctcccccaa gaaaaaacgc
     7141 aaggtggaag atcctaagaa aaagcggaaa gtggacggca ttggtagtgg gagcaacggc
     7201 agcagcggat ccagcgagct gattaaggag aacatgcaca tgaagctgta catggagggc
     7261 accgtggaca accatcactt caagtgcaca tccgagggcg aaggcaagcc ctacgagggc
     7321 acccagacca tgagaatcaa ggtggtcgag ggcggccctc tccccttcgc cttcgacatc
     7381 ctggctacta gcttcctcta cggcagcaag accttcatca accacaccca gggcatcccc
     7441 gacttcttca agcagtcctt ccctgagggc ttcacatggg agagagtcac cacatacgaa
     7501 gacgggggcg tgctgaccgc tacccaggac accagcctcc aggacggctg cctcatctac
     7561 aacgtcaaga tcagaggggt gaacttcaca tccaacggcc ctgtgatgca gaagaaaaca
     7621 ctcggctggg aggccttcac cgagacgctg taccccgctg acggcggcct ggaaggcaga
     7681 aacgacatgg ccctgaagct cgtgggcggg agccatctga tcgcaaacat caagaccaca
     7741 tatagatcca agaaacccgc taagaacctc aagatgcctg gcgtctacta tgtggactac
     7801 agactggaaa gaatcaagga ggccaacaac gagacctacg tcgagcagca cgaggtggca
     7861 gtggccagat actgcgacct ccctagcaaa ctggggcaca agcttaatgg tggaggaggt
     7921 tctggaggcg gtggaagtgg tggcggaggt agcgtcgacc tgcggaccct ggatgtgttt
     7981 tctggctgcg gggggttgtc ggagggattc caccaagcag gcatctctga cacgctgtgg
     8041 gccatcgaga tgtgggaccc tgcggcccag gcgttccggc tgaacaaccc cggctccaca
     8101 gtgttcacag aggactgcaa catcctgctg aagctggtca tggctgggga gaccaccaac
     8161 tcccgcggcc agcggctgcc ccagaaggga gacgtggaga tgctgtgcgg cgggccgccc
     8221 tgccagggct tcagcggcat gaaccgcttc aattcgcgca cctactccaa gttcaaaaac
     8281 tctctggtgg tttccttcct cagctactgc gactactacc ggccccggtt cttcctcctg
     8341 gagaatgtca ggaactttgt ctccttcaag cgctccatgg tcctgaagct caccctccgc
     8401 tgcctggtcc gcatgggcta tcagtgcacc ttcggcgtgc tgcaggccgg tcagtacggc
     8461 gtggcccaga ctaggaggcg ggccatcatc ctggccgcgg cccctggaga gaagctccct
     8521 ctgttcccgg agccactgca cgtgtttgct ccccgggcct gccagctgag cgtggtggtg
     8581 gatgacaaga agtttgtgag caacataacc aggttgagct cgggtccttt ccggaccatc
     8641 acggtgcgag acacgatgtc cgacctgccg gaggtgcgga atggagcctc ggcactggag
     8701 atctcctaca acggggagcc tcagtcctgg ttccagaggc agctccgggg cgcacagtac
     8761 cagcccatcc tcagggacca catctgtaag gacatgagtg cattggtggc tgcccgcatg
     8821 cggcacatcc ccttggcccc agggtcagac tggcgcgatc tgcccaacat cgaggtgcgg
     8881 ctctcagacg gcaccatggc caggaagctg cggtataccc accatgacag gaagaacggc
     8941 cgcagcagct ctggggccct ccgtggggtc tgctcctgcg tggaagccgg caaagcctgc
     9001 gaccccgcag ccaggcagtt caacaccctc atcccctggt gcctgcccca caccgggaac
     9061 cggcacaacc actgggctgg cctctatgga aggctcgagt gggacggctt cttcagcaca
     9121 accgtcacca accccgagcc catgggcaag cagggccgcg tgctccaccc agagcagcac
     9181 cgtgtggtga gcgtgcggga gtgtgcccgc tcccagggct tccctgacac ctaccggctc
     9241 ttcggcaaca tcctggacaa gcaccggcag gtgggcaatg ccgtgccacc gcccctggcc
     9301 aaagccattg gcttggagat caagctttgt atgttggcca aagcccgaga gagtgcctca
     9361 gctaaaataa aggaggagga agctgctaag gactagtaaa tgcatggccg gccttaatta
     9421 aacgcgtctc gagggaattc cgataatcaa cctctggatt acaaaatttg tgaaagattg
     9481 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     9541 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     9601 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     9661 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     9721 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     9781 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaag
     9841 ctgacgtcct ttccatggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     9901 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     9961 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
    10021 gccgcctccc cgcatcggga attcgagctc ggtaccttta agaccaatga cttacaaggc
    10081 agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc taattcactc
    10141 ccaacgaaga caagatctgc tttttgcttg tactgggtct ctctggttag accagatctg
    10201 agcctgggag ctctctggct aactagggaa cccactgctt aagcctcaat aaagcttgcc
    10261 ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac tctggtaact agagatccct
    10321 cagacccttt tagtcagtgt ggaaaatctc tagcagcatc tagctagaat taattccgtg
    10381 tattctatag tgtcacctaa atcgtatgtg tatgatacat aaggttatgt attaattgta
    10441 gccgcgttct aacgacaata tgtacaagcc taattgtgta gcatctggct tactgaagca
    10501 gaccctatca tctctctcgt aaactgccgt cagagtcggt ttggttggac gaaccttctg
    10561 agtttctggt aacgccgtcc cgcacccgga aatggtcagc gaaccaatca gcagggtcat
    10621 cgctagccta gggacgtacc caattcgccc tatagtgagt cgtattacgc gcgctcactg
    10681 gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact taatcgcctt
    10741 gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac cgatcgccct
    10801 tcccaacagt tgcgcagcct gaatggcgaa tgggacgcgc cctgtagcgg cgcattaagc
    10861 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc
    10921 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
    10981 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
    11041 aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
    11101 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
    11161 ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat
    11221 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
    11281 cttacaattt aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
    11341 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
    11401 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
    11461 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
    11521 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
    11581 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
    11641 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
    11701 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
    11761 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
    11821 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
    11881 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
    11941 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
    12001 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
    12061 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
    12121 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
    12181 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
    12241 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
    12301 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
    12361 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
    12421 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
    12481 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
    12541 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
    12601 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
    12661 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
    12721 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
    12781 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
    12841 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
    12901 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
    12961 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
    13021 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
    13081 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
    13141 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
    13201 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
    13261 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
    13321 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
    13381 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
    13441 accatgatta cgccaagcgc gcaattaacc ctcactaaag ggaacaaaag ctggagctgc
//
