LOCUS       sequence-308911-       13706 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid supports Cas9 expression in both males and females, 
            Opie2-dsRed tagged, and can be integrated with pBac or phC31..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13706)
  AUTHORS   Kandul NP, Liu J, Bennett JB, Marshall JM, Akbari OS
  TITLE     A confinable home and rescue gene drive for population modification.
  JOURNAL   Elife. 2021 Mar 5;10. pii: 65939. doi: 10.7554/eLife.65939.
  PUBMED    33666174
REFERENCE   2  (bases 1 to 13706)
  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: "Elife."; 
            date: "2021-03-5"; volume: "10. pii: 65939. doi"; pages: " 
            10.7554/eLife.65939"
COMMENT     Sequence Label: 1095=Rcd-1r-NLS-hSpCas9-T2A-GFP, Opie2-dsRed-SV40
FEATURES             Location/Qualifiers
     source          1..13706
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   651..685
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(866..882)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(866..882)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(879..901)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    890..906
                     /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(914..920,921..938,939..944))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    959..980
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1097..1114)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1268..1856)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1348..1367)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2027..2818,2819..2887))
                     /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     2650..2669
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2888..2992)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3060..3078
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(3116..3138)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     3238..3257
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3451..3473
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3465..3482
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3466..3482
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   4186..4248
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    5282..5351
                     /label=attB
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attB site for the phi-C31 integrase (Groth et al., 
                     2000)"
     CDS             6500..6565
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             6572..6592
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6617..10717
                     /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             6617..10717
                     /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             10718..10765
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             10766..10819
                     /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(10820..10822,10823..10825,10826..11539)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTSKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(11126..11145)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     11473..11494
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             join(12774..12776,12777..12779,12780..13454)
                     /codon_start=1
                     /product="wild-type DsRed"
                     /label=DsRed1
                     /note="mammalian codon-optimized"
                     /translation="MVRSSKNVIKEFMRFKVRMEGTVNGHEFEIEGEGEGRPYEGHNTV
                     KLKVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     VVTVTQDSSLQDGCFIYKVKFIGVNFPSDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     KALKLKDGGHYLVEFKSIYMAKKPVQLPGYYYVDSKLDITSHNEDYTIVEQYERTEGRH
                     HLFL"
     primer_bind     complement(12957..12977)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     13175..13194
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     13369..13392
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    13568..13689
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(13605..13624)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     13659..13678
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
ORIGIN
        1 gatctcggat ctgacaatgt tcagtgcaga gactcggcta cgcctcgtgg actttgaagt
       61 tgaccaacaa tgtttattct tacctctaat agtcctctgt ggcaaggtca agattctgtt
      121 agaagccaat gaagaacctg gttgttcaat aacattttgt tcgtctaata tttcactacc
      181 gcttgacgtt ggctgcactt catgtacctc atctataaac gcttcttctg tatcgctctg
      241 gacgtcatct tcacttacgt gatctgatat ttcactgtca gaatcctcac caacaagctc
      301 gtcatcgctt tgcagaagag cagagaggat atgctcatcg tctaaagaac tacccatttt
      361 attatatatt agtcacgata tctataacaa gaaaatatat atataataag ttatcacgta
      421 agtagaacat gaaataacaa tataattatc gtatgagtta aatcttaaaa gtcacgtaaa
      481 agataatcat gcgtcatttt gactcacgcg gtcgttatag ttcaaaatca gtgacactta
      541 ccgcattgac aagcacgcct cacgggagct ccaagcggcg actgagatgt cctaaatgca
      601 cagcgacgga ttcgcgctat ttagaaagag agagcaatat ttcaagaatg catgcgtcaa
      661 ttttacgcag actatctttc tagggttaaa aaagatttgc gctttactcg acctaaactt
      721 taaacacgtc atagaatctt cgtttgacaa aaaccacatt gtggccaagc tgtgtgacgc
      781 gacgcgcgct aaagaatggc aaaccaagtc gcgcgagcgt cgactctaga ggatccccgg
      841 gtaccgagct cgaattcgta atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc
      901 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
      961 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     1021 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     1081 ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag
     1141 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
     1201 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
     1261 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
     1321 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
     1381 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt
     1441 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     1501 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     1561 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1621 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     1681 ggtggcctaa ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc
     1741 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     1801 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     1861 atcctttgat cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga
     1921 ttttggtcat gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa
     1981 gttttaaatc aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa
     2041 tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc
     2101 ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga
     2161 taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa
     2221 gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt
     2281 gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg
     2341 ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc
     2401 aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg
     2461 gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag
     2521 cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt
     2581 actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt
     2641 caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac
     2701 gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac
     2761 ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag
     2821 caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa
     2881 tactcatact cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga
     2941 gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc
     3001 cccgaaaagt gccacctgac gtctaagaaa ccattattat catgacatta acctataaaa
     3061 ataggcgtat cacgaggccc tttcgtctcg cgcgtttcgg tgatgacggt gaaaacctct
     3121 gacacatgca gctcccggag acggtcacag cttgtctgta agcggatgcc gggagcagac
     3181 aagcccgtca gggcgcgtca gcgggtgttg gcgggtgtcg gggctggctt aactatgcgg
     3241 catcagagca gattgtactg agagtgcacc atatgcggtg tgaaataccg cacagatgcg
     3301 taaggagaaa ataccgcatc aggcgccatt cgccattcag gctgcgcaac tgttgggaag
     3361 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     3421 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     3481 gtgccaagct ttgtttaaaa atataacaaa attgtgatcc cacaaaatga agtggggcaa
     3541 aatcaaataa ttaatagtgt ccgtaaactt gttggtcttc aactttttga ggaacacgtt
     3601 ggacggcaaa tccgtgacta taacacaagt tgatttaata attttagcca acacgtcggg
     3661 ctgcgtgttt tttgccgacg cgtctgtgta cacgttgatt aactggtcga ttaaactgtt
     3721 gaaataattt aatttttggt tcttctttaa atctgtgatg aaatttttta aaataacttt
     3781 aaattcttca ttggtaaaaa atgccacgtt ttgcaacttg tgagggtcta atatgaggtc
     3841 aaactcagta ggagttttat ccaaaaaaga aaacatgatt acgtctgtac acgaacgcgt
     3901 attaacgcag agtgcaaagt ataagagggt taaaaaatat attttacgca ccatatacgc
     3961 atcgggttga tatcgttaat atggatcaat ttgaacagtt gattaacgtg tctctgctca
     4021 agtctttgat caaaacgcaa atcgacgaaa atgtgtcgga caatatcaag tcgatgagcg
     4081 aaaaactaaa aaggctagaa tacgacaatc tcacagacag cgttgagata tacggtattc
     4141 acgacagcag gctgaataat aaaaaaatta gaaactatta tttaacccta gaaagataat
     4201 catattgtga cgtacgttaa agataatcat gcgtaaaatt gacgcatgtg ttttatcggt
     4261 ctgtatatcg aggtttattt attaatttga atagatatta agttttatta tatttacact
     4321 tacatactaa taataaattc aacaaacaat ttatttatgt ttatttattt attaaaaaaa
     4381 aacaaaaact caaaatttct tctataaagt aacaaaactt ttaaacattc tctcttttac
     4441 aaaaataaac ttattttgta ctttaaaaac agtcatgttg tattataaaa taagtaatta
     4501 gcttaactta tacataatag aaacaaatta tacttattag tcagtcagaa acaactttgg
     4561 cacatatcaa tattatgctc tcgacaaata acttttttgc attttttgca cgatgcattt
     4621 gcctttcgcc ttattttaga ggggcagtaa gtacagtaag tacgtttttt cattactggc
     4681 tcttcagtac tgtcatctga tgtaccaggc acttcatttg gcaaaatatt agagatatta
     4741 tcgcgcaaat atctcttcaa agtaggagct tctaaacgct tacgcataaa cgatgacgtc
     4801 aggctcatgt aaaggtttct cataaatttt ttgcgacttt gaaccttttc tcccttgcta
     4861 ctgacattat ggctgtatat aataaaagaa tttatgcagg caatgtttat cattccgtac
     4921 aataatgcca taggccacct attcgtcttc ctactgcagg tcatcacaga acacatttgg
     4981 tctagcgtgt ccactccgcc tttagtttga ttataataca taaccatttg cggtttaccg
     5041 gtactttcgt tgatagaagc atcctcatca caagatgata ataagtatac catcttagct
     5101 ggcttcggtt tatatgagac gagagtaagg ggtccgtcaa aacaaaacat cgatgttccc
     5161 actggcctgg agcgactgtt tttcagtact tccggtatct cgcgtttgtt tgatcgcacg
     5221 gttcccacaa tggttaattc gagctcgccc ggggtcctag gtcgacgatg taggtcacgg
     5281 tctcgaagcc gcggtgcggg tgccagggcg tgcccttggg ctccccgggc gcgtactcca
     5341 cctcacccat ctggtccatc atgatgaacg ggtcgaggtg gcggtagttg atcccggcga
     5401 acgcgcggcg caccgggaag ccctcgccct cgaaaccgct gggcgcggtg gtcacggtga
     5461 gcacgggacg tgcgacggcg tcggcgggtg cggatacgcg gggcagcgtc agcgggttct
     5521 cgacggtcac ggcgggcatg tcgacgcggc cgcgcagaca tagacgagca acaatcaatc
     5581 aattaaatgc ctgaatgcag ctccaaacaa accgagccac gataaccata acatgaccca
     5641 ccggataatg ttcggcatgc aaaagtgaga ttgactttta ggcagacggc aacactatta
     5701 aagatttatt cttgtttctc ggctcgttcg aatctccgat tttctctggg atagttggcc
     5761 aatttatgtg acggactgaa atttttaaag ctctctgcaa caaacaaaat tgaaatgttg
     5821 cacgatgtgg caaagaatac attttctgtt aaccatttct gtattggcaa gtaattccag
     5881 aaatggacaa ctattccaag catccatctg tttaatgggt taaatgcgag ataaaatatt
     5941 tgaaaatttt aaaggcaaat atttgtgtgt gctagaatga tttttgattt atagtatttg
     6001 tcaatacgaa ttcattaatt ctaaaactgg tgtccaaaag tacttaatgg tcattaagcc
     6061 ataaatcgga cctccatcag atgcgcaatt agtctggcca tttcagagag ctaagctatg
     6121 ctatgacata cttaatagaa tcataaattt taatcatcta ccgattaata cttgtaaaat
     6181 atagagaaaa cgtgaatttc cagacgaatt ccaagctgcc aatcaaatca atggtgataa
     6241 gaaacaaaat tctgggagac attcgaacaa gaaattattt attaatttat ggcattcgtg
     6301 cgttcctcgg ctctggccac attgttcggc cacgtgcttt cggtcctttc tcgcttgaag
     6361 cttatatcga taaccacaat agacgccaac tcgaaaccac gtttctcaat cgtccgtcga
     6421 aattccggag ccagatcttt ccaagctact agatctttgc aagctaacgc gaaaaaaaac
     6481 cagctttgcg ctcgagatgg actataagga ccacgacgga gactacaagg atcatgatat
     6541 tgattacaaa gacgatgacg ataagatggc cccaaagaag aagcggaagg tcggtatcca
     6601 cggagtccca gcagccgaca agaagtacag catcggcctg gacatcggca ccaactctgt
     6661 gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg
     6721 caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg
     6781 cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa
     6841 gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag
     6901 cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca
     6961 ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta
     7021 ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct
     7081 ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc
     7141 cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt
     7201 cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact
     7261 gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg
     7321 cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt
     7381 cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga
     7441 caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct
     7501 gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc
     7561 cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa
     7621 agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa
     7681 gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat
     7741 caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga
     7801 ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct
     7861 gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa
     7921 ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc
     7981 caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg
     8041 gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac
     8101 caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga
     8161 gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa
     8221 gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa
     8281 ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcttcga
     8341 ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca cataccacga
     8401 tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg aggacattct
     8461 ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg aggaacggct
     8521 gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc ggcggagata
     8581 caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca agcagtccgg
     8641 caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact tcatgcagct
     8701 gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg tgtccggcca
     8761 gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca ttaagaaggg
     8821 catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc
     8881 cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg gacagaagaa
     8941 cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca gccagatcct
     9001 gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc tgtactacct
     9061 gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc tgtccgacta
     9121 cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg acaacaaggt
     9181 gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg aagaggtcgt
     9241 gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta cccagagaaa
     9301 gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata aggccggctt
     9361 catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac agatcctgga
     9421 ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag tgaaagtgat
     9481 caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt acaaagtgcg
     9541 cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg tgggaaccgc
     9601 cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact acaaggtgta
     9661 cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta
     9721 cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg ccaacggcga
     9781 gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa
     9841 gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa
     9901 aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca agaggaacag
     9961 cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg gcttcgacag
    10021 ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca agtccaagaa
    10081 actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca gcttcgagaa
    10141 gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat
    10201 caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa tgctggcctc
    10261 tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg tgaacttcct
    10321 gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg agcagaaaca
    10381 gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga tcagcgagtt
    10441 ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa
    10501 gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt ttaccctgac
    10561 caatctggga gcccctgccg ccttcaagta ctttgacacc accatcgacc ggaagaggta
    10621 caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca ccggcctgta
    10681 cgagacacgg atcgacctgt ctcagctggg aggcgacaaa aggccggcgg ccacgaaaaa
    10741 ggccggccag gcaaaaaaga aaaaggaggg cagaggaagt cttctaacat gcggtgacgt
    10801 ggaggagaat cccggcccta tggtgagcaa gggcgaggag ctgttcaccg gggtggtgcc
    10861 catcctggtc gagctggacg gcgacgtaaa cggccacaag ttcagcgtgt ccggcgaggg
    10921 cgagggcgat gccacctacg gcaagctgac cctgaagttc atctgcacca ccagcaagct
    10981 gcccgtgccc tggcccaccc tcgtgaccac cctgacctac ggcgtgcagt gcttcagccg
    11041 ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg aaggctacgt
    11101 ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg ccgaggtgaa
    11161 gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact tcaaggagga
    11221 cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg tctatatcat
    11281 ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca acatcgagga
    11341 cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg acggccccgt
    11401 gctgctgccc gacaaccact acctgagcac ccagtccgcc ctgagcaaag accccaacga
    11461 gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca ctctcggcat
    11521 ggacgagctg tacaagtaat taattaacta gaatgaatcg tttttaaaat aacaaatcaa
    11581 ttgttttata atattcgtac gattctttga ttatgtaata aaatgtgatc attaggaaga
    11641 ttacgaaaaa tataaaaaat atgagttctg tgtgtataac aaatgctgta aacgccacaa
    11701 ttgtgtttgt tgcaaataaa cccatgatta tttgattaaa attgttgttt tctttgttca
    11761 tagacaatag tgtgttttgc ctaaacgtgt actgcataaa ctccatgcga gtgtatagcg
    11821 agctagtggc taacgcttgc cccaccaaag tagattcgtc aaaatcctca atttcatcac
    11881 cctcctccaa gtttaacatt tggccgtcgg aattaacttc taaagatgcc acataatcta
    11941 ataaatgaaa tagagattca aacgtggcgt catcgtccgt ttcgaccatt tccgaaaaga
    12001 actcgggcat aaactctatg atttctctgg acgtggtgtt gtcgaaactc tcaaagtacg
    12061 cagtcaggaa cgtgcgcgac atgtcgtcgg gaaactcgcg cggaaacatg ttgttgtaac
    12121 cgaacgggtc ccatagcgcc aaaaccaaat ctgccagcgt caatagaatg agcacgatgc
    12181 cgacaatgga gctggcttgg atagcgattc gagttaacgg ccggggcgcg cccatactcg
    12241 gtggcctccc caccaccaac ttttttgcac tgcaaaaaaa cacgcttttg cacgcgggcc
    12301 catacatagt acaaactcta cgtttcgtag actattttac ataaatagtc tacaccgttg
    12361 tatacgctcc aaatacacta ccacacattg aacctttttg cagtgcaaaa aagtacgtgt
    12421 cggcagtcac gtaggccggc cttatcgggt cgcgtcctgt cacgtacgaa tcacattatc
    12481 ggaccggacg agtgttgtct tatcgtgaca ggacgccagc ttcctgtgtt gctaaccgca
    12541 gccggacgca actccttatc ggaacaggac gcgcctccat atcagccgcg cgttatctca
    12601 tgcgcgtgac cggacacgag gcgcccgtcc cgcttatcgc gcctataaat acagcccgca
    12661 acgatctggt aaacacagtt gaacagcatc tgttacagcg acacaacatg agccggtcca
    12721 acaacgccaa cgcgcccacg ccatccaacc gccgccgcaa cctgtctctg gtgatggtgc
    12781 gctcctccaa gaacgtcatc aaggagttca tgcgcttcaa ggtgcgcatg gagggcaccg
    12841 tgaacggcca cgagttcgag atcgagggcg agggcgaggg ccgcccctac gagggccaca
    12901 acaccgtgaa gctgaaggtg accaagggcg gccccctgcc cttcgcctgg gacatcctgt
    12961 ccccccagtt ccagtacggc tccaaggtgt acgtgaagca ccccgccgac atccccgact
    13021 acaagaagct gtccttcccc gagggcttca agtgggagcg cgtgatgaac ttcgaggacg
    13081 gcggcgtggt gaccgtgacc caggactcct ccctgcagga cggctgcttc atctacaagg
    13141 tgaagttcat cggcgtgaac ttcccctccg acggccccgt aatgcagaag aagaccatgg
    13201 gctgggaggc ctccaccgag cgcctgtacc cccgcgacgg cgtgctgaag ggcgagatcc
    13261 acaaggccct gaagctgaag gacggcggcc actacctggt ggagttcaag tccatctaca
    13321 tggccaagaa gcccgtgcag ctgcccggct actactacgt ggactccaag ctggacatca
    13381 cctcccacaa cgaggactac accatcgtgg agcagtacga gcgcaccgag ggccgccacc
    13441 acctgttcct gtaggactct agatcataat cagccatacc acatttgtag aggttttact
    13501 tgctttaaaa aacctcccac acctccccct gaacctgaaa cataaaatga atgcaattgt
    13561 tgttgttaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
    13621 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
    13681 tgtatcttaa agcttctagg ccggcc
//
