LOCUS       sequence-449615-       14167 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Human expression vector containing SFFV promoter, dCas9 that is 
            fused to 2x NLS, tagBFP and a KRAB domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14167)
  AUTHORS   Gilbert LA, Larson MH, Morsut L, Liu Z, Brar GA, Torres SE, 
            Stern-Ginossar N, Brandman O, Whitehead EH, Doudna JA, Lim WA, 
            Weissman JS, Qi LS
  TITLE     CRISPR-Mediated Modular RNA-Guided Regulation of Transcription in 
            Eukaryotes.
  JOURNAL   Cell. 2013 Jul 9. pii: S0092-8674(13)00826-X. doi: 
            10.1016/j.cell.2013.06.044.
  PUBMED    23849981
REFERENCE   2  (bases 1 to 14167)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Cell. 2013 
            Jul 9. pii: S0092-8674(13)00826-X. doi: 10.1016/j.cell.2013.06.044."
COMMENT     Sequence Label: pHR-SFFV-dCas9-BFP-KRAB
FEATURES             Location/Qualifiers
     source          1..14167
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     253..273
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     primer_bind     complement(343..360)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(514..1102)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(594..613)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1273..2064,2065..2133))
                     /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     1896..1915
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2134..2238)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2306..2324
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2362..2384)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2484..2503
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2717..2736
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     promoter        3026..3044
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     3026..3043
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     LTR             3069..3302
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     3392..3408
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3393..3409
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3412..4000
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3518..3529
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     3927..3947
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4064..4249)
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEP"
     CDS             complement(4295..4990)
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="SELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKVV
                     EGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTAT
                     QDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALKL
                     VGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYCD
                     LPSKLGHKLN"
     CDS             complement(5030..5050)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(5057..5077)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(5090..5116)
                     /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             complement(5096..5122)
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             complement(join(5126..6709,6710..6712,6713..9199,9200..9202,
                     9203..9229))
                     /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             complement(5126..9229)
                     /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"
     regulatory      9226..9235
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(9304..9711)
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     misc_feature    9864..9981
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(10266..10310)
                     /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    10495..10728
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    11225..11350
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             11397..12030
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     polyA_signal    12198..12332
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12222..12241)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     12276..12295
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(12757..12777)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     intron          12907..12972
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     promoter        complement(join(14106..14167,1..268))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(14120..14167,1..88)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
ORIGIN
        1 aggcggcctc ggcctctgca taaataaaaa aaattagtca gccatggggc ggagaatggg
       61 cggaactggg cggagttagg ggcgggatgg gcggagttag gggcgggact atggttgctg
      121 actaattgag atgcatgctt tgcatacttc tgcctgctgg ggagcctggg gactttccac
      181 acctggttgc tgactaattg agatgcatgc tttgcatact tctgcctgct ggggagcctg
      241 gggactttcc acaccctaac tgacacacat tccacagctg cattaatgaa tcggccaacg
      301 cgcggggaga ggcggtttgc gtattgggcg ctcttccgct tcctcgctca ctgactcgct
      361 gcgctcggtc gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt
      421 atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc
      481 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
      541 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
      601 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
      661 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
      721 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
      781 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
      841 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
      901 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
      961 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     1021 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     1081 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     1141 gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac
     1201 ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac
     1261 ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt
     1321 tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt
     1381 accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt
     1441 atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc
     1501 cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa
     1561 tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg
     1621 tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt
     1681 gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc
     1741 agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt
     1801 aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg
     1861 gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac
     1921 tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc
     1981 gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt
     2041 tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg
     2101 aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat attattgaag
     2161 catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa
     2221 acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtct aagaaaccat
     2281 tattatcatg acattaacct ataaaaatag gcgtatcacg aggccctttc gtctcgcgcg
     2341 tttcggtgat gacggtgaaa acctctgaca catgcagctc ccggagacgg tcacagcttg
     2401 tctgtaagcg gatgccggga gcagacaagc ccgtcagggc gcgtcagcgg gtgttggcgg
     2461 gtgtcggggc tggcttaact atgcggcatc agagcagatt gtactgagag tgcaccattc
     2521 gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt
     2581 tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac agtcccccgg
     2641 ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg aagtggcgag
     2701 cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc
     2761 cggtgatgcc ggccacgatg cgtccggcgt agaggatctg gctagcgatg accctgctga
     2821 ttggttcgct gaccatttcc gggtgcggga cggcgttacc agaaactcag aaggttcgtc
     2881 caaccaaacc gactctgacg gcagtttacg agagagatga tagggtctgc ttcagtaagc
     2941 cagatgctac acaattaggc ttgtacatat tgtcgttaga acgcggctac aattaataca
     3001 taaccttatg tatcatacac atacgattta ggtgacacta tagaatacac ggaattaatt
     3061 ctagatgctg ctagagattt tccacactga ctaaaagggt ctgagggatc tctagttacc
     3121 agagtcacac aacagacggg cacacactac ttgaagcact caaggcaagc tttattgagg
     3181 cttaagcagt gggttcccta gttagccaga gagctcccag gctcagatct ggtctaacca
     3241 gagagaccca gtacaagcaa aaagcagatc ttgtcttcgt tgggagtgaa ttagcccttc
     3301 cagtcccccc ttttctttta aaaagtggct aagatctaca gctgccttgt aagtcattgg
     3361 tcttaaaggt accgagctcg aattaattcc ctcgaggtcg acggtatcga tgcggggagg
     3421 cggcccaaag ggagatccga ctcgtctgag ggcgaaggcg aagacgcgga agaggccgca
     3481 gagccggcag caggccgcgg gaaggaaggt ccgctggatt gagggccgaa gggacgtagc
     3541 agaaggacgt cccgcgcaga atccaggtgg caacacaggc gagcagccaa ggaaaggacg
     3601 atgatttccc cgacaacacc acggaattgt cagtgcccaa cagccgagcc cctgtccagc
     3661 agcgggcaag gcaggcggcg atgagttccg ccgtggcaat agggaggggg aaagcgaaag
     3721 tcccggaaag gagctgacag gtggtggcaa tgccccaacc agtgggggtt gcgtcagcaa
     3781 acacagtgca caccacgcca cgttgcctga caacgggcca caactcctca taaagagaca
     3841 gcaaccagga tttatacaag gaggagaaaa tgaaagccat acgggaagca atagcatgat
     3901 acaaaggcat taaagcagcg tatccacata gcgtaaaagg agcaacatag ttaagaatac
     3961 cagtcaatct ttcacaaatt ttgtaatcca gaggttgatt atcgataagc ttgatatcaa
     4021 gcttgcatgc ctgcaggtcg actctagagt cgcggcctta ctagggctct tctcccttct
     4081 ccaaccggag gatcacatct ggcttagtaa gctgataacc caaggaaacc aggttcttat
     4141 agttctccag catcacattt ctgtacacga tctgctgagc agtgtccagc agcttccact
     4201 cctccctggt gaagtccaca aatacatcct tgaaggtcac cagtgtccgg gaccaggcag
     4261 ttagtgactt agcatccatc cctccgccac cgccattaag cttgtgcccc agtttgctag
     4321 ggaggtcgca gtatctggcc actgccacct cgtgctgctc gacgtaggtc tcgttgttgg
     4381 cctccttgat tctttccagt ctgtagtcca catagtagac gccaggcatc ttgaggttct
     4441 tagcgggttt cttggatcta tatgtggtct tgatgtttgc gatcagatgg ctcccgccca
     4501 cgagcttcag ggccatgtcg tttctgcctt ccaggccgcc gtcagcgggg tacagcgtct
     4561 cggtgaaggc ctcccagccg agtgttttct tctgcatcac agggccgttg gatgtgaagt
     4621 tcacccctct gatcttgacg ttgtagatga ggcagccgtc ctggaggctg gtgtcctggg
     4681 tagcggtcag cacgcccccg tcttcgtatg tggtgactct ctcccatgtg aagccctcag
     4741 ggaaggactg cttgaagaag tcggggatgc cctgggtgtg gttgatgaag gtcttgctgc
     4801 cgtagaggaa gctagtagcc aggatgtcga aggcgaaggg gagagggccg ccctcgacca
     4861 ccttgattct catggtctgg gtgccctcgt agggcttgcc ttcgccctcg gatgtgcact
     4921 tgaagtgatg gttgtccacg gtgccctcca tgtacagctt catgtgcatg ttctccttaa
     4981 tcagctcgct ggatccgctg ctgccgttgc tcccactacc aatgccgtcc actttccgct
     5041 ttttcttagg atcttccacc ttgcgttttt tcttggggga gccgctgccc aggctggcat
     5101 aatcgggcac gtcataggga taggcgtcgc ctcccagctg agacaggtcg atccgtgtct
     5161 cgtacaggcc ggtgatgctc tggtggatca gggtggcgtc cagcacctct ttggtgctgg
     5221 tgtacctctt ccggtcgatg gtggtgtcaa agtacttgaa ggcggcaggg gctcccagat
     5281 tggtcagggt aaacaggtgg atgatattct cggcctgctc tctgataggc ttgtctctgt
     5341 gcttgttgta ggcgctcagc accttgtcca gattagcgtc ggccaggatc actctcttgg
     5401 agaactcgct gatctgctcg atgatctcgt ccaggtagtg tttgtgctgt tccacaaaca
     5461 gctgtttctg ctcattatcc tcgggggagc ccttcagctt ctcatagtgg ctggccaggt
     5521 acaggaagtt cacatatttg gagggcaggg ccagttcgtt tcccttctgc agttcgccgg
     5581 cagaggccag cattctcttc cggccgtttt ccagctcgaa cagggagtac ttaggcagct
     5641 tgatgatcag gtcctttttc acttctttgt agcccttggc ttccagaaag tcgatgggat
     5701 tcttctcgaa gctgcttctt tccatgatgg tgatccccag cagctctttc acactcttca
     5761 gtttcttgga cttgcccttt tccactttgg ccaccaccag cacagaatag gccacggtgg
     5821 ggctgtcgaa gccgccgtac ttcttagggt cccagtcctt ctttctggcg atcagcttgt
     5881 cgctgttcct cttgggcagg atagactctt tgctgaagcc gcctgtctgc acctcggtct
     5941 ttttcacgat attcacttgg ggcatagaca gcactttccg cacggtggca aagtcccggc
     6001 ccttatccca cacgatctcg cctgtttcgc cgtttgtctc gatcagaggc cgcttccgga
     6061 tctcgccgtt ggccagggta atctcggtct tgaaaaagtt catgatgttg ctgtagaaga
     6121 agtacttggc ggtagccttg ccgatttcct gctcgctctt ggcgatcatc ttccgcacgt
     6181 cgtacacctt gtagtcgccg tacacgaact cgctttccag cttagggtac tttttgatca
     6241 gggcggttcc cacgacggcg ttcaggtagg cgtcgtgggc gtggtggtag ttgttgatct
     6301 cgcgcacttt gtaaaactgg aaatccttcc ggaaatcgga caccagcttg gacttcaggg
     6361 tgatcacttt cacttcccgg atcagtttgt cgttctcgtc gtacttagtg ttcatccggg
     6421 agtccaggat ctgtgccacg tgctttgtga tctgccgggt ttccaccagc tgtctcttga
     6481 tgaagccggc cttatccagt tcgctcaggc cgcctctctc ggccttggtc agattgtcga
     6541 acttcctctg ggtaatcagc ttggcattca gcagctggcg ccagtagttc ttcatcttct
     6601 tcacgacctc ttcggagggc acgttgtcgc tcttgccccg gttcttgtcg ctccgagtca
     6661 gcactttgtt atcgatggag tcgtccttca gaaagctctg aggcacgata gcgtccacat
     6721 cgtagtcgga cagccggttg atgtccagtt cctggtccac gtacatatcc cgcccattct
     6781 gcaggtagta caggtacagc ttctcgttct gcagctgggt gttttccacg gggtgttctt
     6841 tcaggatctg gctgcccagc tctttgatgc cctcttcgat ccgcttcatt ctctcgcggc
     6901 tgttcttctg tcccttctgg gtggtctggt tctctctggc catttcgatc acgatgttct
     6961 cgggcttgtg ccggcccatc actttcacga gctcgtccac caccttcact gtctgcagga
     7021 tgcccttctt aatggcgggg ctgccggcca gattggcaat gtgctcgtgc aggctatcgc
     7081 cctggccgga cacctgggct ttctggatgt cctctttaaa ggtcaggctg tcgtcgtgga
     7141 tcagctgcat gaagtttctg ttggcgaagc cgtcggactt caggaaatcc aggattgtct
     7201 tgccggactg cttgtcccgg atgccgttga tcagcttccg gctcagcctg ccccagccgg
     7261 tgtatctccg ccgcttcagc tgcttcatca ctttgtcgtc gaacaggtgg gcataggttt
     7321 tcagccgttc ctcgatcatc tctctgtcct caaacagtgt cagggtcagc acgatatctt
     7381 ccagaatgtc ctcgttttcc tcattgtcca ggaagtcctt gtccttgata attttcagca
     7441 gatcgtggta tgtgcccagg gaggcgttga accgatcttc cacgccggag atttccacgg
     7501 agtcgaagca ctcgattttc ttgaagtagt cctctttcag ctgcttcacg gtcactttcc
     7561 ggttggtctt gaacagcagg tccacgatgg cttttttctg ctcgccgctc aggaaggcgg
     7621 gctttctcat tccctcggtc acgtatttca ctttggtcag ctcgttgtac acggtgaagt
     7681 actcgtacag caggctgtgc ttgggcagca ccttctcgtt gggcaggttc ttatcgaagt
     7741 tggtcatccg ctcgatgaag ctctgggcgc tggcgccctt gtccaccact tcctcgaagt
     7801 tccagggggt gatggtttcc tcgctctttc tggtcatcca ggcgaatctg ctgtttcccc
     7861 tggccagagg gcccacgtag taggggatgc ggaaggtcag gatcttctcg atcttttccc
     7921 ggttgtcctt caggaatggg taaaaatctt cctgccgccg cagaatggcg tgcagctctc
     7981 ccaggtggat ctggtggggg atgctgccgt tgtcgaaggt ccgctgcttc cgcagcaggt
     8041 cctctctgtt cagcttcacg agcagttcct cggtgccgtc catcttttcc aggatgggct
     8101 tgatgaactt gtagaactct tcctggctgg ctccgccatc gatgtagccg gcgtagccgt
     8161 tcttgctctg gtcgaagaaa atctctttgt acttctcagg cagctgctgc cgcacgagag
     8221 ctttcagcag ggtcaggtcc tggtggtgct cgtcgtatct cttgatcata gaggcgctca
     8281 ggggggcctt ggtgatctcg gtgttcactc tcaggatgtc gctcagcagg atggcgtcgg
     8341 acaggttctt ggcggccaga aacaggtcgg cgtactggtc gccgatctgg gccagcaggt
     8401 tgtccaggtc gtcgtcgtag gtgtccttgc tcagctgcag tttggcatcc tcggccaggt
     8461 cgaagttgct cttgaagttg ggggtcaggc ccaggctcag ggcaatcagg ttgccgaaca
     8521 ggccattctt cttctcgccg ggcagctggg cgatcagatt ttccagccgt ctgctcttgc
     8581 tcagtctggc agacaggatg gccttggcgt ccacgccgct ggcgttgatg gggttttcct
     8641 cgaacagctg gttgtaggtc tgcaccagct ggatgaacag cttgtccacg tcgctgttgt
     8701 cggggttcag gtcgccctcg atcaggaagt ggccccggaa cttgatcatg tgggccaggg
     8761 ccagatagat cagccgcagg tcggccttgt cggtgctgtc caccagtttc tttctcaggt
     8821 ggtagatggt ggggtacttc tcgtggtagg ccacctcgtc cacgatgttg ccgaagatgg
     8881 ggtgccgctc gtgcttctta tcctcttcca ccaggaagga ctcttccagt ctgtggaaga
     8941 agctgtcgtc caccttggcc atctcgttgc tgaagatctc ttgcagatag cagatccggt
     9001 tcttccgtct ggtgtatctt cttctggcgg ttctcttcag ccgggtggcc tcggctgttt
     9061 ctccgctgtc gaacagcagg gcgccgatca ggttcttctt gatgctgtgc cggtcggtgt
     9121 tgcccagcac cttgaatttc ttgctgggca ccttgtactc gtcggtgatc acggcccagc
     9181 ccacagagtt ggtgccgatg gccaggccga tgctgtactt cttgtccatg gtggccgggc
     9241 ccgcggtacc gcaagcttga tatcctgcag acgcgtgaga attctcgaga gctccagatc
     9301 ccccccgggc gactcagtct gtcggaggac tggcgcgccg agtgaggggt tgtgagctct
     9361 tttatagagc tcgggaagca gaagcgcgcg aacagaagcg agaagcaggc tgattggtta
     9421 attcaaataa ggcgcagggt catttcaggt ccttggggga gcctggaaac atctgatggg
     9481 tcttaagaaa ctgctgaggg ttgggccata tctggggacc atctgttctt ggcctcgggc
     9541 cggggccgaa actgcggtga ccatctgttc ttggccccgg gccggggccg aaactgctca
     9601 ccgcagatat cctgtttggc ccaacgttag ctattttcat gtacccgccc ttgatctgaa
     9661 cttctctatt cttggtttgg tatttttcca tgccttgcaa aatggcgtta ctgcagctag
     9721 cttgccaaac ctacaggtgg ggtctttcat tccccccttt ttctggagac taaataaaat
     9781 cttttatttt atcggggctg caggaattcg atatcaagct tatcgatcca aactggatct
     9841 ctgctgtccc tgtaataaac ccgaaaattt tgaatttttg taatttgttt ttgtaattct
     9901 ttagtttgta tgtctgttgc tattatgtct actattcttt cccctgcact gtacccccca
     9961 atcccccctt ttcttttaaa attgtggatg aatactgcca tttggcgata ccgtcgagat
    10021 ccgttcacta atcgaatgga tctgtctctg tctctctctc caccttcttc ttctattcct
    10081 tcgggcctgt cgggtcccct cggggttggg aggtgggtct gaaacgataa tggtgaatat
    10141 ccctgcctaa ctctattcac tatagaaagt acagcaaaaa ctattcttaa acctaccaag
    10201 cctcctacta tcattatgaa taattttata taccacagcc aatttgttat gttaaaccaa
    10261 ttccacaaac ttgcccattt atctaattcc aataattctt gttcattctt ttcttgctgg
    10321 ttttgcgatt cttcaattaa ggagtgtatt aagcttgtgt aattgttaat ttctctgtcc
    10381 cactccatcc aggtcgtgtg attccaaatc tgttccagag atttattact ccaactagca
    10441 ttccaaggca cagcagtggt gcaaatgagt tttccagagc aaccccaaat ccccaggagc
    10501 tgttgatcct ttaggtatct ttccacagcc aggattcttg cctggagctg cttgatgccc
    10561 cagactgtga gttgcaacag atgctgttgc gcctcaatag ccctcagcaa attgttctgc
    10621 tgctgcacta taccagacaa taattgtctg gcctgtaccg tcagcgtcat tgacgctgcg
    10681 cccatagtgc ttcctgctgc tcccaagaac ccaaggaaca aagctcctat tcccactgct
    10741 cttttttctc tctgcaccac tcttctcttt gccttggtgg gtgctactcc taatggttca
    10801 atttttacta ctttatattt atataattca cttctccaat tgtccctcat atctcctcct
    10861 ccaggtctga agatcagcgg ccggccgctt gctgtgcggt ggtcttactt ttgttttgct
    10921 cttcctctat cttgtctaaa gcttccttgg tgtcttttat ctctatcctt tgatgcacac
    10981 aatagagggt tgctactgta ttatataatg atctaagttc ttctgatcct gtctgaaggg
    11041 atggttgtag ctgtcccagt atttgtctac agccttctga tgtttctaac aggccaggat
    11101 taactgcgaa tcgttctagc tccctgcttg cccatactat atgttttaat ttatattttt
    11161 tctttccccc tggccttaac cgaatttttt cccatcgcga tctaattctc ccccgcttaa
    11221 tactgacgct ctcgcaccca tctctctcct tctagcctcc gctagtcaaa atttttggcg
    11281 tactcaccag tcgccgcccc tcgcctcttg ccgtgcgcgc ttcagcaagc cgagtcctgc
    11341 gtcgagagag ctcctctggt ttccctttcg ctttcaagtc cctgttcggg cgccactgct
    11401 agagattttc cacactgact aaaagggtct gagggatctc tagttaccag agtcacacaa
    11461 cagacgggca cacactactt gaagcactca aggcaagctt tattgaggct taagcagtgg
    11521 gttccctagt tagccagaga gctcccaggc tcagatctgg tctaaccaga gagacccagt
    11581 acaggcaaaa agcagctgct tatatgcagg atctgagggc tcgccactcc ccagtcccgc
    11641 ccaggccacg cctccctgga aagtccccag cggaaagtcc cttgtagcaa gctcgatatc
    11701 agcagttctt gaagtactcc ggatgcagct ctcgggccac gtgatgaaat gctaggcggc
    11761 tgtcaaacct ccactctaac acttctctct ccgggtcatc catcccatgc aggctcacag
    11821 ggtgtaacaa gctggtgttc tctcctttat tggcctcttc taccttatct ggctcaactg
    11881 gtactagctt gtagcaccat ccaaaggtca gtggatatct gacccctggc cctggtgtgt
    11941 agttctgcta atcagggaag tagccttgtg tgtggtagat ccacagatca aggatatctt
    12001 gtcttctttg ggagtgaatt agcccttcca actactaagt ttgtagtaca tatttaacaa
    12061 atacaatttc tttaaaatga aaataattca gaggaatcac aggtttagag taaatgaaac
    12121 cacaggtaat tggcagtggt aatagggtat ggggtgggaa gtttgggatg attttggtta
    12181 gcttgagtta tccagttgat ccagacatga taagatacat tgatgagttt ggacaaacca
    12241 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
    12301 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
    12361 ttcaggttca gggggaggtg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
    12421 gtatggctga ttatgatctc tagtcaaggc actatacatc aaatattcct tattaacccc
    12481 tttacaaatt aaaaagctaa aggtacacaa tttttgagca tagttattaa tagcagacac
    12541 tctatgcctg tgtggagtaa gaaaaaacag tatgttatga ttataactgt tatgcctact
    12601 tataaaggtt acagaatatt tttccataat tttcttgtat agcagtgcag ctttttcctt
    12661 tgtggtgtaa atagcaaagc aagcaagagt tctattacta aacacagcat gactcaaaaa
    12721 acttagcaat tctgaaggaa agtccttggg gtcttctacc tttctcttct tttttggagg
    12781 agtagaatgt tgagagtcag cagtagcctc atcatcacta gatggcattt cttctgagca
    12841 aaacaggttt tcctcattaa aggcattcca ccactgctcc cattcatcag ttccataggt
    12901 tggaatctaa aatacacaaa caattagaat cagtagttta acacattata cacttaaaaa
    12961 ttttatattt accttagagc tttaaatctc tgtaggtagt ttgtccaatt atgtcacacc
    13021 acagaagtaa ggttccttca caaagatccg gggcccactc ataaatccag ttgccgccac
    13081 ggtagccaat caccgtatcg tataaatcat cgtcggtacg ttcggcatcg ctcatcacaa
    13141 tacgtgcctg gacgtcgagg atttcgcgtg ggtcaatgcc gcgccagatc cacatcagac
    13201 ggttaatcat gcgataccag tgagggatgg ttttaccatc aagggccgac tgcacaggcg
    13261 gttgtgcgcc gtgattaaag cggcggacta gcgtcgaggt ttcaggatgt ttaaagcggg
    13321 gtttgaacag ggtttcgctc aggtttgcct gtgtcatgga tgcagcctcc agaatactta
    13381 ctggaaacta ttgtaacccg cctgaagtta aaaagaacaa cgcccggcag tgccaggcgt
    13441 tgaaaagatt agcgaccgga gattggcggg acgaatacga cgcccatatc ccacggctgt
    13501 tcaatccagg tatcttgcgg gatatcaaca acatagtcat caaccagcgg acgaccagcc
    13561 ggttttgcga agatggtgac aaagtgcgct tttggataca tttcacgaat cgcaaccgca
    13621 gtaccaccgg tatccaccag gtcatcaata acgatgaagc cttcgccatc gccttctgcg
    13681 cgtttcagca ctttaagctc gcgctggttg tcgtgatcgt agctggaaat acaaacggta
    13741 tcgacatgac gaatacccag ttcacgcgcc agtaacgcac ccggtacaga ccgccacggc
    13801 ttacggcaat aatgcctttc cattgttcag aaggcatcag tcggcttgcg agtttacgtg
    13861 catggatctg caacatgtcc caggtgacga tgtatttttc gctcatgtga agtgtcccag
    13921 cctgtttatc tacggcttaa aaagtgttcg aggggaaaat aggttgcgcg agattataga
    13981 gatctggcgc actaaaaacc agtatttcac atgagtccgc gtctttttac gcactgcctc
    14041 tccctgacgc gggataaagt ggtattctca aacatatctc gcaagcctgt cttgtgtcca
    14101 agctttttgc aaaagcctag gcctccaaaa aagcctcctc actacttctg gaatagctca
    14161 gaggccg
//
