LOCUS       sequence-209090-       10418 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses dCas9-PmCDA1-UGI and gRNA(HPRT) in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10418)
  AUTHORS   Nishida K, Arazoe T, Yachie N, Banno S, Kakimoto M, Tabata M, 
            Mochizuki M, Miyabe A, Araki M, Hara KY, Shimatani Z, Kondo A
  TITLE     Targeted nucleotide editing using hybrid prokaryotic and vertebrate 
            adaptive immune systems.
  JOURNAL   Science. 2016 Aug 4. pii: aaf8729.
  PUBMED    27492474
REFERENCE   2  (bases 1 to 10418)
  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: "Science. 
            2016 Aug 4. pii: aaf8729."
COMMENT     Sequence Label: pcDNA3.1_pCMV-dCas-PmCDA1-ugi pH1-gRNA(HPRT)
FEATURES             Location/Qualifiers
     source          1..10418
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(35..55)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(35..53)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(74..90)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(74..90)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(87..109)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    98..114
                     /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)."
     promoter        complement(join(122..128,129..146,147..152))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    167..188
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(305..322)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(476..1064)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(556..575)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1235..2026,2027..2095))
                     /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     1858..1877
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2096..2200)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2226..2681)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2363..2384)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2575..2594
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2808..2830
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2822..2839
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2823..2839
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2849..2867
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3027..3046
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     misc_RNA        3136..3211
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        3226..3605
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3606..3809
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3760..3780
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        3854..3872
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             3901..3921
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(3946..3972,3973..3975,3976..8049)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3946..8049
                     /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
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             8062..8082
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8290..8355
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8989..9009
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             9016..9069
                     /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             9240..9260
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             9267..9320
                     /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             9327..10121
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     9991..10010
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    10295..10416
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10332..10351)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10386..10405
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 ggccgccacc gcggtggagc tccagctttt gttcccttta gtgagggtta attgcgcgct
       61 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
      121 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
      181 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
      241 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
      301 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
      361 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
      421 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
      481 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
      541 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
      601 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
      661 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
      721 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
      781 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
      841 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
      901 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
      961 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
     1021 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
     1081 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
     1141 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
     1201 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
     1261 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
     1321 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
     1381 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
     1441 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
     1501 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
     1561 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
     1621 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
     1681 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
     1741 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
     1801 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
     1861 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
     1921 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
     1981 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
     2041 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
     2101 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
     2161 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
     2221 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
     2281 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
     2341 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
     2401 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
     2461 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
     2521 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
     2581 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
     2641 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
     2701 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
     2761 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
     2821 ttgtaaaacg acggccagtg agcgcgcgta atacgactca ctatagggcg aattgggtac
     2881 cgggccaatt cgaacgctga cgtcatcaac ccgctccaag gaatcgcggg cccagtgtca
     2941 ctaggcggga acacccagcg cgcgtgcgcc ctggcaggaa gatggctgtg agggacaggg
     3001 gagtggcgcc ctgcaatatt tgcatgtcgc tatgtgttct gggaaatcac cataaacgtg
     3061 aaatgtcttt ggatttggga atcttataag ttctgtatga ggaccacaga tccccccgag
     3121 atgtcatgaa agagagtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta
     3181 tcaacttgaa aaagtggcac cgagtcggtg ctttttttac gcgttgacat tgattattga
     3241 ctagttatta atagtaatca attacggggt cattagttca tagcccatat atggagttcc
     3301 gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
     3361 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
     3421 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
     3481 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
     3541 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
     3601 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
     3661 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
     3721 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
     3781 tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc actgcttact
     3841 ggcttatcga aattaatacg actcactata gggagaccca agctggctag cgaaatggca
     3901 ccgaagaaga agcgtaaagt cggaatccac ggagttcctg cggcaatgga caagaagtac
     3961 tccattgggc tcgctatcgg cacaaacagc gtcggttggg ccgtcattac ggacgagtac
     4021 aaggtgccga gcaaaaaatt caaagttctg ggcaataccg atcgccacag cataaagaag
     4081 aacctcattg gcgccctcct gttcgactcc ggggagacgg ccgaagccac gcggctcaaa
     4141 agaacagcac ggcgcagata tacccgcaga aagaatcgga tctgctacct gcaggagatc
     4201 tttagtaatg agatggctaa ggtggatgac tctttcttcc ataggctgga ggagtccttt
     4261 ttggtggagg aggataaaaa gcacgagcgc cacccaatct ttggcaatat cgtggacgag
     4321 gtggcgtacc atgaaaagta cccaaccata tatcatctga ggaagaagct tgtagacagt
     4381 actgataagg ctgacttgcg gttgatctat ctcgcgctgg cgcatatgat caaatttcgg
     4441 ggacacttcc tcatcgaggg ggacctgaac ccagacaaca gcgatgtcga caaactcttt
     4501 atccaactgg ttcagactta caatcagctt ttcgaagaga acccgatcaa cgcatccgga
     4561 gttgacgcca aagcaatcct gagcgctagg ctgtccaaat cccggcggct cgaaaacctc
     4621 atcgcacagc tccctgggga gaagaagaac ggcctgtttg gtaatcttat cgccctgtca
     4681 ctcgggctga cccccaactt taaatctaac ttcgacctgg ccgaagatgc caagcttcaa
     4741 ctgagcaaag acacctacga tgatgatctc gacaatctgc tggcccagat cggcgaccag
     4801 tacgcagacc tttttttggc ggcaaagaac ctgtcagacg ccattctgct gagtgatatt
     4861 ctgcgagtga acacggagat caccaaagct ccgctgagcg ctagtatgat caagcgctat
     4921 gatgagcacc accaagactt gactttgctg aaggcccttg tcagacagca actgcctgag
     4981 aagtacaagg aaattttctt cgatcagtct aaaaatggct acgccggata cattgacggc
     5041 ggagcaagcc aggaggaatt ttacaaattt attaagccca tcttggaaaa aatggacggc
     5101 accgaggagc tgctggtaaa gcttaacaga gaagatctgt tgcgcaaaca gcgcactttc
     5161 gacaatggaa gcatccccca ccagattcac ctgggcgaac tgcacgctat cctcaggcgg
     5221 caagaggatt tctacccctt tttgaaagat aacagggaaa agattgagaa aatcctcaca
     5281 tttcggatac cctactatgt aggccccctc gcccggggaa attccagatt cgcgtggatg
     5341 actcgcaaat cagaagagac catcactccc tggaacttcg aggaagtcgt ggataagggg
     5401 gcctctgccc agtccttcat cgaaaggatg actaactttg ataaaaatct gcctaacgaa
     5461 aaggtgcttc ctaaacactc tctgctgtac gagtacttca cagtttataa cgagctcacc
     5521 aaggtcaaat acgtcacaga agggatgaga aagccagcat tcctgtctgg agagcagaag
     5581 aaagctatcg tggacctcct cttcaagacg aaccggaaag ttaccgtgaa acagctcaaa
     5641 gaagactatt tcaaaaagat tgaatgtttc gactctgttg aaatcagcgg agtggaggat
     5701 cgcttcaacg catccctggg aacgtatcac gatctcctga aaatcattaa agacaaggac
     5761 ttcctggaca atgaggagaa cgaggacatt cttgaggaca ttgtcctcac ccttacgttg
     5821 tttgaagata gggagatgat tgaagaacgc ttgaaaactt acgctcatct cttcgacgac
     5881 aaagtcatga aacagctcaa gaggcgccga tatacaggat gggggcggct gtcaagaaaa
     5941 ctgatcaatg ggatccgaga caagcagagt ggaaagacaa tcctggattt tcttaagtcc
     6001 gatggatttg ccaaccggaa cttcatgcag ttgatccatg atgactctct cacctttaag
     6061 gaggacatcc agaaagcaca agtttctggc cagggggaca gtcttcacga gcacatcgct
     6121 aatcttgcag gtagcccagc tatcaaaaag ggaatactgc agaccgttaa ggtcgtggat
     6181 gaactcgtca aagtaatggg aaggcataag cccgagaata tcgttatcga gatggcccga
     6241 gagaaccaaa ctacccagaa gggacagaag aacagtaggg aaaggatgaa gaggattgaa
     6301 gagggtataa aagaactggg gtcccaaatc cttaaggaac acccagttga aaacacccag
     6361 cttcagaatg agaagctcta cctgtactac ctgcagaacg gcagggacat gtacgtggat
     6421 caggaactgg acatcaatcg gctctccgac tacgacgtgg atcatatcgt gccccagtct
     6481 tttctcaaag atgattctat tgataataaa gtgttgacaa gatccgataa aaatagaggg
     6541 aagagtgata acgtcccctc agaagaagtt gtcaagaaaa tgaaaaatta ttggcggcag
     6601 ctgctgaacg ccaaactgat cacacaacgg aagttcgata atctgactaa ggctgaacga
     6661 ggtggcctgt ctgagttgga taaagccggc ttcatcaaaa ggcagcttgt tgagacacgc
     6721 cagatcacca agcacgtggc ccaaattctc gattcacgca tgaacaccaa gtacgatgaa
     6781 aatgacaaac tgattcgaga ggtgaaagtt attactctga agtctaagct ggtctcagat
     6841 ttcagaaagg actttcagtt ttataaggtg agagagatca acaattacca ccatgcgcat
     6901 gatgcctacc tgaatgcagt ggtaggcact gcacttatca aaaaatatcc caagcttgaa
     6961 tctgaatttg tttacggaga ctataaagtg tacgatgtta ggaaaatgat cgcaaagtct
     7021 gagcaggaaa taggcaaggc caccgctaag tacttctttt acagcaatat tatgaatttt
     7081 ttcaagaccg agattacact ggccaatgga gagattcgga agcgaccact tatcgaaaca
     7141 aacggagaaa caggagaaat cgtgtgggac aagggtaggg atttcgcgac agtccggaag
     7201 gtcctgtcca tgccgcaggt gaacatcgtt aaaaagaccg aagtacagac cggaggcttc
     7261 tccaaggaaa gtatcctccc gaaaaggaac agcgacaagc tgatcgcacg caaaaaagat
     7321 tgggacccca agaaatacgg cggattcgat tctcctacag tcgcttacag tgtactggtt
     7381 gtggccaaag tggagaaagg gaagtctaaa aaactcaaaa gcgtcaagga actgctgggc
     7441 atcacaatca tggagcgatc aagcttcgaa aaaaacccca tcgactttct cgaggcgaaa
     7501 ggatataaag aggtcaaaaa agacctcatc attaagcttc ccaagtactc tctctttgag
     7561 cttgaaaacg gccggaaacg aatgctcgct agtgcgggcg agctgcagaa aggtaacgag
     7621 ctggcactgc cctctaaata cgttaatttc ttgtatctgg ccagccacta tgaaaagctc
     7681 aaagggtctc ccgaagataa tgagcagaag cagctgttcg tggaacaaca caaacactac
     7741 cttgatgaga tcatcgagca aataagcgaa ttctccaaaa gagtgatcct cgccgacgct
     7801 aacctcgata aggtgctttc tgcttacaat aagcacaggg ataagcccat cagggagcag
     7861 gcagaaaaca ttatccactt gtttactctg accaacttgg gcgcgcctgc agccttcaag
     7921 tacttcgaca ccaccataga cagaaagcgg tacacctcta caaaggaggt cctggacgcc
     7981 acactgattc atcagtcaat tacggggctc tatgaaacaa gaatcgacct ctctcagctc
     8041 ggtggagaca gcagggctga ccccaagaag aagaggaagg tgggtggagg aggtaccggc
     8101 ggtggaggct cagcagaata cgtacgagct ctgtttgact tcaatgggaa tgacgaggag
     8161 gatctcccct ttaagaaggg cgatattctc cgcatcagag ataagcccga agaacaatgg
     8221 tggaatgccg aggatagcga agggaaaagg ggcatgattc tggtgccata tgtggagaaa
     8281 tattccggtg actacaaaga ccatgatggg gattacaaag accacgacat cgactacaaa
     8341 gacgacgacg ataaatcagg gatgacagac gccgagtacg tgcgcattca tgagaaactg
     8401 gatatttaca ccttcaagaa gcagttcttc aacaacaaga aatctgtgtc acaccgctgc
     8461 tacgtgctgt ttgagttgaa gcgaaggggc gaaagaaggg cttgcttttg gggctatgcc
     8521 gtcaacaagc cccaaagtgg caccgagaga ggaatacacg ctgagatatt cagtatccga
     8581 aaggtggaag agtatcttcg ggataatcct gggcagttta cgatcaactg gtattccagc
     8641 tggagtcctt gcgctgattg tgccgagaaa attctggaat ggtataatca ggaacttcgg
     8701 ggaaacgggc acacattgaa aatctgggcc tgcaagctgt actacgagaa gaatgcccgg
     8761 aaccagatag gactctggaa tctgagggac aatggtgtag gcctgaacgt gatggtttcc
     8821 gagcactatc agtgttgtcg gaagattttc atccaaagct ctcataacca gctcaatgaa
     8881 aaccgctggt tggagaaaac actgaaacgt gcggagaagt ggagatccga gctgagcatc
     8941 atgatccagg tcaagattct gcataccact aagtctccag ccgttggtcc caagaagaaa
     9001 agaaaagtcg gtaccgaagg caggggaagc cttctgactt gtggggatgt ggaagaaaac
     9061 cctggtccat ctagaatgat tgaacaagat ggattgcacg caggttctcc ggccgcttgg
     9121 gtggagaggc tattcggcta tgacgcggtg tgacamagrg tgcggagaag tggagatccg
     9181 agctgagcat catgatccag gtcaagattc tgcataccac taagtctcca gccgttggtc
     9241 ccaagaagaa aagaaaagtc ggtaccgaag gcaggggaag ccttctgact tgtggggatg
     9301 tggaagaaaa ccctggtcca tctagaatga ttgaacaaga tggattgcac gcaggttctc
     9361 cggccgcttg ggtggagagg ctattcggct atgactgggc acaacagaca atcggctgct
     9421 ctgatgccgc cgtgttccgg ctgtcagcgc aggggcgccc ggttcttttt gtcaagaccg
     9481 acctgtccgg tgccctgaat gaactgcagg acgaggcagc gcggctatcg tggctggcca
     9541 cgacgggcgt tccttgcgca gctgtgctcg acgttgtcac tgaagcggga agggactggc
     9601 tgctattggg cgaagtgccg gggcaggatc tcctgtcatc tcaccttgct cctgccgaga
     9661 aagtatccat catggctgat gcaatgcggc ggctgcatac gcttgatccg gctacctgcc
     9721 cattcgacca ccaagcgaaa catcgcatcg agcgagcacg tactcggatg gaagccggtc
     9781 ttgtcgatca ggatgatctg gacgaagagc atcaggggct cgcgccagcc gaactgttcg
     9841 ccaggctcaa ggcgcgcatg cccgacggcg aggatctcgt cgtgacccat ggcgatgcct
     9901 gcttgccgaa tatcatggtg gaaaatggcc gcttttctgg attcatcgac tgtggccggc
     9961 tgggtgtggc ggaccgctat caggacatag cgttggctac ccgtgatatt gctgaagagc
    10021 ttggcggcga atgggctgac cgcttcctcg tgctttacgg tatcgccgct cccgattcgc
    10081 agcgcatcgc cttctatcgc cttcttgacg agttcttctg agcgggactc tggggttcga
    10141 aatgaccgac caagcgacgc ccaacctgcc atcacgagat ttcgattcca ccgccgcctt
    10201 ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga tcctccagcg
    10261 cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag cttataatgg
    10321 ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt cactgcattc
    10381 tagttgtggt ttgtccaaac tcatcaatgt atcttagc
//
