LOCUS       sequence-438177-       11585 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses dCAS9-SunTag fusion protein and double marker GFP-T2A-Puro
            under independent promoters.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11585)
  TITLE     Systemic comparison of dCas9-based DNA methylation editing systems 
            for specificity and stability
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 11585)
  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: 
            "Unpublished"
COMMENT     Sequence Label: M3-dCAS9-SunTag (dCas9-SunTag)
FEATURES             Location/Qualifiers
     source          1..11585
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        178..463
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     misc_RNA        complement(560..635)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(654..902)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(712..731)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(882..902)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     rep_origin      complement(964..1552)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1044..1063)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1723..2514,2515..2583))
                     /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     2346..2365
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2584..2688)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2756..2774
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2812..2834)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2934..2953
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(2970..3425)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3107..3128)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3319..3338
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   3500..3640
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3511..3640
                     /label=AAV2 ITR
     polyA_signal    3654..3775
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3665..3684)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3719..3738
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(3943..4542)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     4523..4542
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     CDS             complement(4543..4596)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(join(4627..5337,5338..5340,5341..5343))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(4627..5337,5338..5340,5341..5343))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4669..4690)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     5018..5037
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     regulatory      5340..5349
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(5388..5591)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(5417..5437)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        5592..5895
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     polyA_signal    5910..6117
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6106..6123
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(6173..6193)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(6218..6274)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6290..6346)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6362..6418)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6434..6490)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6506..6562)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6578..6634)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6650..6706)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6722..6778)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6794..6850)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6866..6922)
                     /codon_start=1
                     /product="GCN4 peptide optimized to improve solubility 
                     while preserving binding to an scFv-GFP fusion protein 
                     (Tanenbaum et al., 2014)"
                     /label=GCN4_v4
                     /translation="EELLSKNYHLENEVARLKK"
     CDS             complement(6944..6964)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(6971..7018)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(join(7019..8602,8603..8605,8606..11092,
                     11093..11095,11096..11119))
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(7019..11119)
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(11144..11164)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(11171..11236)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     regulatory      11236..11245
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     intron          11257..11484
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     promoter        complement(join(11485..11585,1..176))
                     /label=chicken beta-actin promoter
ORIGIN
        1 ctctccgcct cgccccgccc cgcccctcgc cccgccccgc cccgcctggc gcgcgccccc
       61 cccccccccc gcccccatcg ctgcacaaaa taattaaaaa ataaataaat acaaaattgg
      121 gggtggggag gggggggaga tggggagagt gaagcagaac gtggggctca cctcgaccat
      181 ggtaatagcg atgactaata cgtagatgta ctgccaagta ggaaagtccc ataaggtcat
      241 gtactgggca caatgccagg cgggccattt accgtcattg acgtcaatag ggggcgtact
      301 tggcatatga tacacttgat gtactgccaa gtgggcagtt taccgtaaat actccaccca
      361 ttgacgtcaa tggaaagtcc ctattggcgt tactattgac gtcaatgggc gggggtcgtt
      421 gggcggtcag ccaggcgggc catttaccgt aagttatgta acgggtacct ctagagccat
      481 ttgtctgcag aattggcgca cgcgctaaaa acggactagc cttattttaa cttgctattt
      541 ctagctctaa aacaaaaaag caccgactcg gtgccacttt ttcaagttga taacggacta
      601 gccttatttt aacttgctat ttctagctct aaaacaggtc ttctcgaaga cccggtgttt
      661 cgtcctttcc acaagatata taaagccaag aaatcgaaat actttcaagt tacggtaagc
      721 atatgatagt ccattttaaa acataatttt aaaactgcaa actacccaag aaattattac
      781 tttctacgtc acgtattttg tactaatatc tttgtgttta cagtcaaatt aattccaatt
      841 atctctctaa cagccttgta tcgtatatgc aaatatgaag gaatcatggg aaataggccc
      901 tcacatgtga gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc
      961 gtttttccat aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag
     1021 gtggcgaaac ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt
     1081 gcgctctcct gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg
     1141 aagcgtggcg ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg
     1201 ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg
     1261 taactatcgt cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac
     1321 tggtaacagg attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg
     1381 gcctaactac ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt
     1441 taccttcgga aaaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg
     1501 tggttttttt gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc
     1561 tttgatcttt tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     1621 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     1681 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttaccaat gcttaatcag
     1741 tgaggcacct atctcagcga tctgtctatt tcgttcatcc atagttgcct gactccccgt
     1801 cgtgtagata actacgatac gggagggctt accatctggc cccagtgctg caatgatacc
     1861 gcggcttcca cgctcaccgg ctccagattt atcagcaata aaccagccag ccggaagggc
     1921 cgagcgcaga agtggtcctg caactttatc cgcctccatc cagtctatta attgttgccg
     1981 ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc aacgttgttg ccattgctac
     2041 aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca ttcagctccg gttcccaacg
     2101 atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc
     2161 tccgatcgtt gtcagaagta agttggccgc agtgttatca ctcatggtta tggcagcact
     2221 gcataattct cttactgtca tgccatccgt aagatgcttt tctgtgactg gtgagtactc
     2281 aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat
     2341 acgggataat accgcgccac atagcagaac tttaaaagtg ctcatcattg gaaaacgttc
     2401 ttcggggcga aaactctcaa ggatcttacc gctgttgaga tccagttcga tgtaacccac
     2461 tcgtgcaccc aactgatctt cagcatcttt tactttcacc agcgtttctg ggtgagcaaa
     2521 aacaggaagg caaaatgccg caaaaaaggg aataagggcg acacggaaat gttgaatact
     2581 catactcttc ctttttcaat attattgaag catttatcag ggttattgtc tcatgagcgg
     2641 atacatattt gaatgtattt agaaaaataa acaaataggg gttccgcgca catttccccg
     2701 aaaagtgcca cctgacgtct aagaaaccat tattatcatg acattaacct ataaaaatag
     2761 gcgtatcacg aggccctttc gtctcgcgcg tttcggtgat gacggtgaaa acctctgaca
     2821 catgcagctc ccggagacgg tcacagcttg tctgtaagcg gatgccggga gcagacaagc
     2881 ccgtcagggc gcgtcagcgg gtgttggcgg gtgtcggggc tggcttaact atgcggcatc
     2941 agagcagatt gtactgagag tgcaccataa aattgtaaac gttaatattt tgttaaaatt
     3001 cgcgttaaat ttttgttaaa tcagctcatt ttttaaccaa tagaccgaaa tcggcaaaat
     3061 cccttataaa tcaaaagaat agcccgagat agagttgagt gttgttccag tttggaacaa
     3121 gagtccacta ttaaagaacg tggactccaa cgtcaaaggg cgaaaaaccg tctatcaggg
     3181 cgatggccca ctacgtgaac catcacccaa atcaagtttt ttggggtcga ggtgccgtaa
     3241 agcactaaat cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc
     3301 gaacgtggcg agaaaggaag ggaagaaagc gaaaggagcg ggcgctaagg cgctggcaag
     3361 tgtagcggtc acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg
     3421 cgcgtactat ggttgctttg acgtatgcgg tgtgaaatac cgcacagatg cgtaaggaga
     3481 aaataccgca tcaggcgccc ctgcaggcag ctgcgcgctc gctcgctcac tgaggccgcc
     3541 cgggcaaagc ccgggcgtcg ggcgaccttt ggtcgcccgg cctcagtgag cgagcgagcg
     3601 cgcagagagg gagtggccaa ctccatcact aggggttcct gcggccgccg cgttaagata
     3661 cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct ttatttgtga
     3721 aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac aagttaacaa
     3781 caacaattgc attcatttta tgtttcaggt tcagggggag gtgtgggagg ttttttaaag
     3841 caagtaaaac ctctacaaat gtggtatggc tgattatgat cagttatcta gatccggtgg
     3901 atcccgggcc cgcggtaccg tcgactgcag aattcgaagc tttcaggcac cgggcttgcg
     3961 ggtcatgcac caggtgcgcg gtccttcggg cacctcgacg tcggcggtga cggtgaagcc
     4021 gagccgctcg tagaagggga ggttgtgggg cgcggaggtc tccaggaagg cgggcacccc
     4081 ggcgcgctcg gccgcctcca ctccggggag cacgacggcg ctgcccagac ccttgccctg
     4141 gtggtcgggc gagacgccga cggtggccag gaaccacgcg ggctccttgg gccggtgcgg
     4201 cgccaggagg ccttccatct gttgctgcgc ggccagccgg gaaccgctca actcggccat
     4261 gcgcgggccg atctcggcga acaccgcccc cgcttcgacg ctctccggcg tggtccagac
     4321 cgccaccgcg gcgccgtcgt ccgcgaccca caccttgccg atgtcgagcc cgacgcgcgt
     4381 gaggaagagt tcttgcagct cggtgacccg ctcgatgtgg cggtccggat cgacggtgtg
     4441 gcgcgtggcg gggtagtcgg cgaacgcggc ggcgagggtg cgtacggccc tggggacgtc
     4501 gtcgcgggtg gcgaggcgca ccgtgggctt gtactcggtc attgggccag gattctcctc
     4561 gacgtcaccg catgttagca gacttcctct gccctcagaa gcttgagctc gagatctgag
     4621 tccggacttg tacagctcgt ccatgccgag agtgatcccg gcggcggtca cgaactccag
     4681 caggaccatg tgatcgcgct tctcgttggg gtctttgctc agggcggact gggtgctcag
     4741 gtagtggttg tcgggcagca gcacggggcc gtcgccgatg ggggtgttct gctggtagtg
     4801 gtcggcgagc tgcacgctgc cgtcctcgat gttgtggcgg atcttgaagt tcaccttgat
     4861 gccgttcttc tgcttgtcgg ccatgatata gacgttgtgg ctgttgtagt tgtactccag
     4921 cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg cccttcagct cgatgcggtt
     4981 caccagggtg tcgccctcga acttcacctc ggcgcgggtc ttgtagttgc cgtcgtcctt
     5041 gaagaagatg gtgcgctcct ggacgtagcc ttcgggcatg gcggacttga agaagtcgtg
     5101 ctgcttcatg tggtcggggt agcggctgaa gcactgcacg ccgtaggtca gggtggtcac
     5161 gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag atgaacttca gggtcagctt
     5221 gccgtaggtg gcatcgccct cgccctcgcc ggacacgctg aacttgtggc cgtttacgtc
     5281 gccgtccagc tcgaccagga tgggcaccac cccggtgaac agctcctcgc ccttgctcac
     5341 catggtggcg accggtagcg ctagcggatc tgacggttca ctaaaccagc tctgcttata
     5401 tagacctccc accgtacacg cctaccgccc atttgcgtca atggggcgga gttgttacga
     5461 cattttggaa agtcccgttg attttggtgc caaaacaaac tcccattgac gtcaatgggg
     5521 tggagacttg gaaatccccg tgagtcaaac cgctatccac gcccattgat gtactgccaa
     5581 aaccgcatca ccatggtaat agcgatgact aatacgtaga tgtactgcca agtaggaaag
     5641 tcccataagg tcatgtactg ggcacaatgc caggcgggcc atttaccgtc attgacgtca
     5701 atagggggcg tacttggcat atgatacact tgatgtactg ccaagtgggc agtttaccgt
     5761 aaatactcca cccattgacg tcaatggaaa gtccctattg gcgttactat gggaacatac
     5821 gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc agccaggcgg gccatttacc
     5881 gtaagttatg taacgcggaa cgcggccgct ccccagcatg cctgctattc tcttcccaat
     5941 cctccccctt gctgtcctgc cccaccccac cccccagaat agaatgacac ctactcagac
     6001 aatgcgatgc aatttcctca ttttattagg aaaggacagt gggagtggca ccttccaggg
     6061 tcaaggaagg cacgggggag gggcaaacaa cagatggctg gcaactagaa ggcacagtcg
     6121 aggctgatca gcgagctcta gttagaattg aattcgggcc ggcccctcat ccgaccttac
     6181 gtttcttctt cggactacct cctccggaac caccgccctt tttgagccta gcaacttcgt
     6241 tctcaagatg ataatttttc gatagcaatt cctcgccaga accactcccc ttttttagtc
     6301 tcgctacctc attttccaga tgataatttt tactcagtaa ttcttcaccc gaccctgaac
     6361 ctttctttaa tcgagctact tcgttttcga ggtggtaatt ctttgaaagt agttcctcac
     6421 cactgcccga tccctttttt aatcgagcga cctcattttc aagatgatag ttcttcgaga
     6481 gcagttcctc tcctgagccg gaacccttct tcaaacgcgc tacctcgttc tcaagatgat
     6541 aattcttgga tagtagctct tcaccactgc cacttccctt ttttaagcgt gccacctcat
     6601 tttcgaggtg gtagttttta gataatagct cttctccaga accactgccc ttttttagcc
     6661 gagccacttc gttttcgaga tgataatttt tcgagagtaa ctcctcaccc gatccggacc
     6721 ctttctttag tcgtgctact tcgttctcca aatgataatt tttgctcagc aactcctccc
     6781 cacttccgct cccctttttc agtctagcta cctcattttc caggtggtaa ttctttgaaa
     6841 gcagttcttc tccactgcca gaacctttct taagacgagc cacttcgttc tcaagatgat
     6901 aattcttgct caaaagttct tcgcttccgc cgccagcgcg tcccactttt ctttttttct
     6961 taggctcgag ctttttcttt ttggcctgtc cggccttttt cgtggccgcc ggccttttgt
     7021 cgcctcccag ctgagacagg tcgatccgtg tctcgtacag gccggtgatg ctctggtgga
     7081 tcagggtggc gtccagcacc tctttggtgc tggtgtacct cttccggtcg atggtggtgt
     7141 caaagtactt gaaggcggca ggggctccca gattggtcag ggtaaacagg tggatgatat
     7201 tctcggcctg ctctctgatg ggcttatccc ggtgcttgtt gtaggcggac agcactttgt
     7261 ccagattagc gtcggccagg atcactctct tggagaactc gctgatctgc tcgatgatct
     7321 cgtccaggta gtgcttgtgc tgttccacaa acagctgttt ctgctcatta tcctcggggg
     7381 agcccttcag cttctcatag tggctggcca ggtacaggaa gttcacatat ttggagggca
     7441 gggccagttc gtttcccttc tgcagttcgc cggcagaggc cagcattctc ttccggccgt
     7501 tttccagctc gaacagggag tacttaggca gcttgatgat caggtccttt ttcacttctt
     7561 tgtagccctt ggcttccaga aagtcgatgg gattcttctc gaagctgctt ctttccatga
     7621 tggtgatccc cagcagctct ttcacactct tcagtttctt ggacttgccc ttttccactt
     7681 tggccaccac cagcacagaa taggccacgg tggggctgtc gaagccgccg tacttcttag
     7741 ggtcccagtc cttctttctg gcgatcagct tatcgctgtt cctcttgggc aggatagact
     7801 ctttgctgaa gccgcctgtc tgcacctcgg tctttttcac gatattcact tggggcatgc
     7861 tcagcacttt ccgcacggtg gcaaaatccc ggcccttatc ccacacgatc tccccggttt
     7921 cgccgtttgt ctcgatcaga ggccgcttcc ggatctcgcc gttggccagg gtaatctcgg
     7981 tcttgaaaaa gttcatgatg ttgctgtaga agaagtactt ggcggtagcc ttgccgattt
     8041 cctgctcgct cttggcgatc atcttccgca cgtcgtacac cttgtagtcg ccgtacacga
     8101 actcgctttc cagcttaggg tactttttga tcagggcggt tcccacgacg gcgttcaggt
     8161 aggcgtcgtg ggcgtggtgg tagttgttga tctcgcgcac tttgtaaaac tggaaatcct
     8221 tccggaaatc ggacaccagc ttggacttca gggtgatcac tttcacttcc cggatcagct
     8281 tgtcattctc gtcgtactta gtgttcatcc gggagtccag gatctgtgcc acgtgctttg
     8341 tgatctgccg ggtttccacc agctgtctct tgatgaagcc ggccttatcc agttcgctca
     8401 ggccgcctct ctcggccttg gtcagattgt cgaactttct ctgggtaatc agcttggcgt
     8461 tcagcagctg ccgccagtag ttcttcatct tcttcacgac ctcttcggag ggcacgttgt
     8521 cgctcttgcc ccggttcttg tcgcttctgg tcagcacctt gttgtcgatg gagtcgtcct
     8581 tcagaaagct ctgaggcacg atagcgtcca catcgtagtc ggacagccgg ttgatgtcca
     8641 gttcctggtc cacgtacata tcccgcccat tctgcaggta gtacaggtac agcttctcgt
     8701 tctgcagctg ggtgttttcc acggggtgtt ctttcaggat ctggctgccc agctctttga
     8761 tgccctcttc gatccgcttc attctctcgc ggctgttctt ctgtcccttc tgggtggtct
     8821 ggttctctct ggccatttcg atcacgatgt tctcgggctt gtgccggccc atcactttca
     8881 cgagctcgtc caccaccttc actgtctgca ggatgccctt cttaatggcg gggctgccgg
     8941 ccagattggc aatgtgctcg tgcaggctat cgccctggcc ggacacctgg gctttctgga
     9001 tgtcctcttt aaaggtcagg ctgtcgtcgt ggatcagctg catgaagttt ctgttggcga
     9061 agccgtcgga cttcaggaaa tccaggattg tcttgccgga ctgcttgtcc cggatgccgt
     9121 tgatcagctt ccggctcagc ctgccccagc cggtgtatct ccgccgcttc agctgcttca
     9181 tcactttgtc gtcgaacagg tgggcatagg ttttcagccg ttcctcgatc atctctctgt
     9241 cctcaaacag tgtcagggtc agcacgatat cttccagaat gtcctcgttt tcctcattgt
     9301 ccaggaagtc cttgtccttg ataattttca gcagatcgtg gtatgtgccc agggaggcgt
     9361 tgaaccgatc ttccacgccg gagatttcca cggagtcgaa gcactcgatt ttcttgaagt
     9421 agtcctcttt cagctgcttc acggtcactt tccggttggt cttgaacagc aggtccacga
     9481 tggccttttt ctgctcgccg ctcaggaagg cgggctttct cattccctcg gtcacgtatt
     9541 tcactttggt cagctcgtta tacacggtga agtactcgta cagcaggctg tgcttgggca
     9601 gcaccttctc gttgggcagg ttcttatcga agttggtcat ccgctcgatg aagctctggg
     9661 cggaagcgcc cttgtccacc acttcctcga agttccaggg ggtgatggtt tcctcgctct
     9721 ttctggtcat ccaggcgaat ctgctgtttc ccctggccag agggcccacg tagtagggga
     9781 tgcggaaggt caggatcttc tcgatctttt cccggttgtc cttcaggaat gggtaaaaat
     9841 cttcctgccg ccgcagaatg gcgtgcagct ctcccaggtg gatctggtgg gggatgctgc
     9901 cgttgtcgaa ggtccgctgc ttccgcagca ggtcctctct gttcagcttc acgagcagtt
     9961 cctcggtgcc gtccatcttt tccaggatgg gcttgatgaa cttgtagaac tcttcctggc
    10021 tggctccgcc gtcaatgtag ccggcgtagc cgttcttgct ctggtcgaag aaaatctctt
    10081 tgtacttctc aggcagctgc tgccgcacga gagctttcag cagggtcagg tcctggtggt
    10141 gctcgtcgta tctcttgatc atagaggcgc tcaggggggc cttggtgatc tcggtgttca
    10201 ctctcaggat gtcgctcagc aggatggcgt cggacaggtt cttggcggcc agaaacaggt
    10261 cggcgtactg gtcgccgatc tgggccagca ggttgtccag gtcgtcgtcg taggtgtcct
    10321 tgctcagctg cagtttggca tcctcggcca ggtcgaagtt gctcttgaag ttgggggtca
    10381 ggcccaggct cagggcaatc aggttgccga acaggccatt cttcttctcg ccgggcagct
    10441 gggcgatcag attttccagc cgtctgctct tgctcagtct ggcagacagg atggccttgg
    10501 cgtccacgcc gctggcgttg atggggtttt cctcgaacag ctggttgtag gtctgcacca
    10561 gctggatgaa cagcttgtcc acgtcgctgt tgtcggggtt caggtcgccc tcgatcagga
    10621 agtggccccg gaacttgatc atgtgggcca gggccagata gatcagccgc aggtcggcct
    10681 tgtcggtgct gtccaccagt ttctttctca ggtggtagat ggtggggtac ttctcgtggt
    10741 aggccacctc gtccacgatg ttgccgaaga tggggtgccg ctcgtgcttc ttatcctctt
    10801 ccaccaggaa ggactcttcc agtctgtgga agaagctgtc gtccaccttg gccatctcgt
    10861 tgctgaagat ctcttgcaga tagcagatcc ggttcttccg tctggtgtat cttcttctgg
    10921 cggttctctt cagccgggtg gcctcggctg tttcgccgct gtcgaacagc agggctccga
    10981 tcaggttctt cttgatgctg tgccggtcgg tgttgcccag caccttgaat ttcttgctgg
    11041 gcaccttgta ctcgtcggtg atcacggccc agcccacaga gttggtgccg atggccaggc
    11101 cgatgctgta cttcttgtcg gctgctggga ctccgtggat accgaccttc cgcttcttct
    11161 ttggggccat cttatcgtca tcgtctttgt aatcaatatc atgatccttg tagtctccgt
    11221 cgtggtcctt atagtccatg gtggcaccgg tccaacctga aaaaaagtga tttcaggcag
    11281 gtgctccagg taattaaaca ttaatacccc accaaccaac catcccttaa acccttacct
    11341 cttgctcagc taattacagc ccggaggaga agggccgtcc cgcccgctca cctgtgggag
    11401 taacgcggtc agtcagagcc ggggcgggcg gcgcgaggcg gcggcggagc ggggcacggg
    11461 gcgaaggcag cgcgcagcga ctcccgcccg ccgcgcgctt cgctttttat agggccgccg
    11521 ccgccgccgc ctcgccataa aaggaaactt tcggagcgcg ccgctctgat tggctgccgc
    11581 cgcac
//
