LOCUS       sequence-333526-       11850 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Delivers all prime editing nuclease components in a single, GFP 
            selectable plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11850)
  AUTHORS   Adikusuma F, Lushington C, Arudkumar J, Godahewa GI, Chey YCJ, 
            Gierus L, Piltz S, Geiger A, Jain Y, Reti D, Wilson LOW, Bauer DC, 
            Thomas PQ
  TITLE     Optimized nickase- and nuclease-based prime editing in human and 
            mouse cells.
  JOURNAL   Nucleic Acids Res. 2021 Sep 17. pii: 6371974. doi: 
            10.1093/nar/gkab792.
  PUBMED    34534334
REFERENCE   2  (bases 1 to 11850)
  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: "Nucleic 
            Acids Res. 2021 Sep 17. pii: 6371974. doi: 10.1093/nar/gkab792."
COMMENT     Sequence Label: PEA1-Nuclease-GFP
FEATURES             Location/Qualifiers
     source          1..11850
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             5521..5568
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5584..5637
                     /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             join(5638..5640,5641..6351)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(5638..5640,5641..6351)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5941..5960)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6288..6309
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(6379..6396)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6385..6592
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     6601..6617
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     6601..6617
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     misc_RNA        complement(6657..6732)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(6750..6998)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     repeat_region   7007..7147
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   7007..7136
                     /label=AAV2 ITR
     rep_origin      7222..7677
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7309..7328)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7519..7540
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7694..7713)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7813..7835
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7873..7891)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7959..8063
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8064..8132,8133..8924)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(8282..8301)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9095..9683
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9584..9603
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        9745..9993
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        10012..10087
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        10172..10457
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        10459..10732
                     /label=chicken beta-actin promoter
     intron          10733..10960
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      10972..10981
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             10981..11046
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             11053..11073
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(11098..11850,1..3348)
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             join(11098..11850,1..3348)
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
ORIGIN
        1 aagagcaact tcgacctggc cgaggatgcc aaactgcagc tgagcaagga cacctacgac
       61 gacgacctgg acaacctgct ggcccagatc ggcgaccagt acgccgacct gtttctggcc
      121 gccaagaacc tgtccgacgc catcctgctg agcgacatcc tgagagtgaa caccgagatc
      181 accaaggccc ccctgagcgc ctctatgatc aagagatacg acgagcacca ccaggacctg
      241 accctgctga aagctctcgt gcggcagcag ctgcctgaga agtacaaaga gattttcttc
      301 gaccagagca agaacggcta cgccggctac attgacggcg gagccagcca ggaagagttc
      361 tacaagttca tcaagcccat cctggaaaag atggacggca ccgaggaact gctcgtgaag
      421 ctgaacagag aggacctgct gcggaagcag cggaccttcg acaacggcag catcccccac
      481 cagatccacc tgggagagct gcacgccatt ctgcggcggc aggaagattt ttacccattc
      541 ctgaaggaca accgggaaaa gatcgagaag atcctgacct tccgcatccc ctactacgtg
      601 ggccctctgg ccaggggaaa cagcagattc gcctggatga ccagaaagag cgaggaaacc
      661 atcaccccct ggaacttcga ggaagtggtg gacaagggcg cttccgccca gagcttcatc
      721 gagcggatga ccaacttcga taagaacctg cccaacgaga aggtgctgcc caagcacagc
      781 ctgctgtacg agtacttcac cgtgtataac gagctgacca aagtgaaata cgtgaccgag
      841 ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa aggccatcgt ggacctgctg
      901 ttcaagacca accggaaagt gaccgtgaag cagctgaaag aggactactt caagaaaatc
      961 gagtgcttcg actccgtgga aatctccggc gtggaagatc ggttcaacgc ctccctgggc
     1021 acataccacg atctgctgaa aattatcaag gacaaggact tcctggacaa tgaggaaaac
     1081 gaggacattc tggaagatat cgtgctgacc ctgacactgt ttgaggacag agagatgatc
     1141 gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa gcagctgaag
     1201 cggcggagat acaccggctg gggcaggctg agccggaagc tgatcaacgg catccgggac
     1261 aagcagtccg gcaagacaat cctggatttc ctgaagtccg acggcttcgc caacagaaac
     1321 ttcatgcagc tgatccacga cgacagcctg acctttaaag aggacatcca gaaagcccag
     1381 gtgtccggcc agggcgatag cctgcacgag cacattgcca atctggccgg cagccccgcc
     1441 attaagaagg gcatcctgca gacagtgaag gtggtggacg agctcgtgaa agtgatgggc
     1501 cggcacaagc ccgagaacat cgtgatcgaa atggccagag agaaccagac cacccagaag
     1561 ggacagaaga acagccgcga gagaatgaag cggatcgaag agggcatcaa agagctgggc
     1621 agccagatcc tgaaagaaca ccccgtggaa aacacccagc tgcagaacga gaagctgtac
     1681 ctgtactacc tgcagaatgg gcgggatatg tacgtggacc aggaactgga catcaaccgg
     1741 ctgtccgact acgatgtgga ccatatcgtg cctcagagct ttctgaagga cgactccatc
     1801 gacaacaagg tgctgaccag aagcgacaag aaccggggca agagcgacaa cgtgccctcc
     1861 gaagaggtcg tgaagaagat gaagaactac tggcggcagc tgctgaacgc caagctgatt
     1921 acccagagaa agttcgacaa tctgaccaag gccgagagag gcggcctgag cgaactggat
     1981 aaggccggct tcatcaagag acagctggtg gaaacccggc agatcacaaa gcacgtggca
     2041 cagatcctgg actcccggat gaacactaag tacgacgaga atgacaagct gatccgggaa
     2101 gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt tccggaagga tttccagttt
     2161 tacaaagtgc gcgagatcaa caactaccac cacgcccacg acgcctacct gaacgccgtc
     2221 gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt gtacggcgac
     2281 tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg agcaggaaat cggcaaggct
     2341 accgccaagt acttcttcta cagcaacatc atgaactttt tcaagaccga gattaccctg
     2401 gccaacggcg agatccggaa gcggcctctg atcgagacaa acggcgaaac cggggagatc
     2461 gtgtgggata agggccggga ttttgccacc gtgcggaaag tgctgagcat gccccaagtg
     2521 aatatcgtga aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc tatcctgccc
     2581 aagaggaaca gcgataagct gatcgccaga aagaaggact gggaccctaa gaagtacggc
     2641 ggcttcgaca gccccaccgt ggcctattct gtgctggtgg tggccaaagt ggaaaagggc
     2701 aagtccaaga aactgaagag tgtgaaagag ctgctgggga tcaccatcat ggaaagaagc
     2761 agcttcgaga agaatcccat cgactttctg gaagccaagg gctacaaaga agtgaaaaag
     2821 gacctgatca tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg ccggaagaga
     2881 atgctggcct ctgccggcga actgcagaag ggaaacgaac tggccctgcc ctccaaatat
     2941 gtgaacttcc tgtacctggc cagccactat gagaagctga agggctcccc cgaggataat
     3001 gagcagaaac agctgtttgt ggaacagcac aagcactacc tggacgagat catcgagcag
     3061 atcagcgagt tctccaagag agtgatcctg gccgacgcta atctggacaa agtgctgtcc
     3121 gcctacaaca agcaccggga taagcccatc agagagcagg ccgagaatat catccacctg
     3181 tttaccctga ccaatctggg agcccctgcc gccttcaagt actttgacac caccatcgac
     3241 cggaagaggt acaccagcac gaaagaggtg ctggacgcca ccctgatcca ccagagcatc
     3301 accggcctgt acgagacacg gatcgacctg tctcagctgg gaggtgactc tggaggatct
     3361 agcggaggat cctctggcag cgagacacca ggaacaagcg agtcagcaac accagagagc
     3421 agtggcggca gcagcggcgg cagcagcacc ctaaatatag aagatgagta tcggctacat
     3481 gagacctcaa aagagccaga tgtttctcta gggtccacat ggctgtctga ttttcctcag
     3541 gcctgggcgg aaaccggggg catgggactg gcagttcgcc aagctcctct gatcatacct
     3601 ctgaaagcaa cctctacccc cgtgtccata aaacaatacc ccatgtcaca agaagccaga
     3661 ctggggatca agccccacat acagagactg ttggaccagg gaatactggt accctgccag
     3721 tccccctgga acacgcccct gctacccgtt aagaaaccag ggactaatga ttataggcct
     3781 gtccaggatc tgagagaagt caacaagcgg gtggaggaca tccaccccac cgtgcccaac
     3841 ccttacaacc tcttgagcgg gctcccaccg tcccaccagt ggtacactgt gcttgattta
     3901 aaggatgcct ttttctgcct gagactccac cccaccagtc agcctctctt cgcctttgag
     3961 tggagagatc cagagatggg aatctcagga caattgacct ggaccagact cccacagggt
     4021 ttcaaaaaca gtcccaccct gtttaatgag gcactgcaca gagacctagc agacttccgg
     4081 atccagcacc cagacttgat cctgctacag tacgtggatg acttactgct ggccgccact
     4141 tctgagctag actgccaaca aggtactcgg gccctgttac aaaccctagg gaacctcggg
     4201 tatcgggcct cggccaagaa agcccaaatt tgccagaaac aggtcaagta tctggggtat
     4261 cttctaaaag agggtcagag atggctgact gaggccagaa aagagactgt gatggggcag
     4321 cctactccaa agacccctcg acaactaagg gagttcctag ggaaggcagg cttctgtcgc
     4381 ctcttcatcc ctgggtttgc agaaatggca gcccccctgt accctctcac caaaccgggg
     4441 actctgttta attggggccc agaccaacaa aaggcctatc aagaaatcaa gcaagccctt
     4501 ctaactgccc cagccctggg gttgccagat ttgactaagc cctttgaact ctttgtcgac
     4561 gagaagcagg gctacgccaa aggtgtccta acgcaaaaac tgggaccttg gcgtcggccg
     4621 gtggcctacc tgtccaaaaa gctagaccca gtagcagctg ggtggccccc ttgcctacgg
     4681 atggtagcag ccattgccgt actgacaaag gatgcaggca agctaaccat gggacagcca
     4741 ctagtcattc tggcccccca tgcagtagag gcactagtca aacaaccccc cgaccgctgg
     4801 ctttccaacg cccggatgac tcactatcag gccttgcttt tggacacgga ccgggtccag
     4861 ttcggaccgg tggtagccct gaacccggct acgctgctcc cactgcctga ggaagggctg
     4921 caacacaact gccttgatat cctggccgaa gcccacggaa cccgacccga cctaacggac
     4981 cagccgctcc cagacgccga ccacacctgg tacacggatg gaagcagtct cttacaagag
     5041 ggacagcgta aggcgggagc tgcggtgacc accgagaccg aggtaatctg ggctaaagcc
     5101 ctgccagccg ggacatccgc tcagcgggct gaactgatag cactcaccca ggccctaaag
     5161 atggcagaag gtaagaagct aaatgtttat actgatagcc gttatgcttt tgctactgcc
     5221 catatccatg gagaaatata cagaaggcgt gggtggctca catcagaagg caaagagatc
     5281 aaaaataaag acgagatctt ggccctacta aaagccctct ttctgcccaa aagacttagc
     5341 ataatccatt gtccaggaca tcaaaaggga cacagcgccg aggctagagg caaccggatg
     5401 gctgaccaag cggcccgaaa ggcagccatc acagagactc cagacacctc taccctcctc
     5461 atagaaaatt catcaccctc tggcggctca aaaagaaccg ccgacggcag cgaattcgag
     5521 aaaaggccgg cggccacgaa aaaggccggc caggcaaaaa agaaaaagga attcggcagt
     5581 ggagagggca gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc tggcccagtg
     5641 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     5701 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     5761 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     5821 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     5881 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     5941 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     6001 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     6061 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     6121 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     6181 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     6241 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     6301 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa ggaattctaa
     6361 ctagagctcg ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc
     6421 cctcccccgt gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa
     6481 atgaggaaat tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg
     6541 ggcaggacag caagggggag gattgggaag agaatagcag gcatgctggg gagcggccgc
     6601 tctagaacta gtggatcccc cgggctgcag gaattcgata atcctgcagg aaaaaagcac
     6661 cgactcggtg ccactttttc aagttgataa cggactagcc ttattttaac ttgctatttc
     6721 tagctctaaa acgtcttctc gaagaccccg gtgtttcgtc ctttccacaa gatatataaa
     6781 gccaagaaat cgaaatactt tcaagttacg gtaagcatat gatagtccat tttaaaacat
     6841 aattttaaaa ctgcaaacta cccaagaaat tattactttc tacgtcacgt attttgtact
     6901 aatatctttg tgtttacagt caaattaatt ccaattatct ctctaacagc cttgtatcgt
     6961 atatgcaaat atgaaggaat catgggaaat aggccctcgc ggccgcagga acccctagtg
     7021 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
     7081 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc
     7141 ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     7201 atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     7261 tggttacgcg cagcgtgacc gctacacttg ccagcgcctt agcgcccgct cctttcgctt
     7321 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     7381 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg
     7441 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     7501 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aactctatct
     7561 cgggctattc ttttgattta taagggattt tgccgatttc ggtctattgg ttaaaaaatg
     7621 agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaattttat
     7681 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
     7741 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
     7801 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
     7861 cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
     7921 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
     7981 ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga taaatgcttc
     8041 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
     8101 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
     8161 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
     8221 agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact tttaaagttc
     8281 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
     8341 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
     8401 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
     8461 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
     8521 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
     8581 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
     8641 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     8701 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     8761 ctggagccgg tgagcgtgga agccgcggta tcattgcagc actggggcca gatggtaagc
     8821 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     8881 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     8941 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     9001 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     9061 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     9121 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     9181 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     9241 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     9301 acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag tcgtgtctta
     9361 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     9421 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     9481 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     9541 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     9601 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     9661 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     9721 tttgctggcc ttttgctcac atgtgagggc ctatttccca tgattccttc atatttgcat
     9781 atacgataca aggctgttag agagataatt ggaattaatt tgactgtaaa cacaaagata
     9841 ttagtacaaa atacgtgacg tagaaagtaa taatttcttg ggtagtttgc agttttaaaa
     9901 ttatgtttta aaatggacta tcatatgctt accgtaactt gaaagtattt cgatttcttg
     9961 gctttatata tcttgtggaa aggacgaaac accgggtctt cgagaagacc tgttttagag
    10021 ctagaaatag caagttaaaa taaggctagt ccgttatcaa cttgaaaaag tggcaccgag
    10081 tcggtgcggg tcttcgagaa gacctttttt ttgttctagt ccgtttttag cgcgtgcgcc
    10141 aattctgcag acaaatggct ctagaggtac ccgttacata acttacggta aatggcccgc
    10201 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat agtaacgcca atagggactt
    10261 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
    10321 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
    10381 attgtgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
    10441 tcatcgctat taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc
    10501 ccccctcccc acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg
    10561 gggcgggggg ggggggcgcg cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc
    10621 gaggcggaga ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt ttccttttat
    10681 ggcgaggcgg cggcggcggc ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc
    10741 tgcgcgctgc cttcgccccg tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct
    10801 ctgactgacc gcgttactcc cacaggtgag cgggcgggac ggcccttctc ctccgggctg
    10861 taattagctg agcaagaggt aagggtttaa gggatggttg gttggtgggg tattaatgtt
    10921 taattacctg gagcacctgc ctgaaatcac tttttttcag gttggaccgg tgccaccatg
    10981 gactataagg accacgacgg agactacaag gatcatgata ttgattacaa agacgatgac
    11041 gataagatgg ccccaaagaa gaagcggaag gtcggtatcc acggagtccc agcagccgac
    11101 aagaagtaca gcatcggcct ggacatcggc accaactctg tgggctgggc cgtgatcacc
    11161 gacgagtaca aggtgcccag caagaaattc aaggtgctgg gcaacaccga ccggcacagc
    11221 atcaagaaga acctgatcgg agccctgctg ttcgacagcg gcgaaacagc cgaggccacc
    11281 cggctgaaga gaaccgccag aagaagatac accagacgga agaaccggat ctgctatctg
    11341 caagagatct tcagcaacga gatggccaag gtggacgaca gcttcttcca cagactggaa
    11401 gagtccttcc tggtggaaga ggataagaag cacgagcggc accccatctt cggcaacatc
    11461 gtggacgagg tggcctacca cgagaagtac cccaccatct accacctgag aaagaaactg
    11521 gtggacagca ccgacaaggc cgacctgcgg ctgatctatc tggccctggc ccacatgatc
    11581 aagttccggg gccacttcct gatcgagggc gacctgaacc ccgacaacag cgacgtggac
    11641 aagctgttca tccagctggt gcagacctac aaccagctgt tcgaggaaaa ccccatcaac
    11701 gccagcggcg tggacgccaa ggccatcctg tctgccagac tgagcaagag cagacggctg
    11761 gaaaatctga tcgcccagct gcccggcgag aagaagaatg gcctgttcgg aaacctgatt
    11821 gccctgagcc tgggcctgac ccccaacttc
//
