LOCUS       sequence-316542-       16657 bp    DNA     circular SYN 14-APR-2025
DEFINITION  Drives expression of Cas9 and a TdTomato fluorescent marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 16657)
  AUTHORS   Gamez S, Chaverra-Rodriguez D, Buchman A, Kandul NP, Mendez-Sanchez 
            SC, Bennett JB, Sanchez C HM, Yang T, Antoshechkin I, Duque JE, 
            Papathanos PA, Marshall JM, Akbari OS
  TITLE     Exploiting a Y chromosome-linked Cas9 for sex selection and gene 
            drive.
  JOURNAL   Nat Commun. 2021 Dec 10;12(1):7202. doi: 10.1038/s41467-021-27333-1.
  PUBMED    34893590
REFERENCE   2  (bases 1 to 16657)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 14, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat 
            Commun."; date: "2021-12-10"; volume: "12(1):7202. doi"; pages: " 
            10.1038/s41467-021-27333-1"
COMMENT     Sequence Label: SGyA
FEATURES             Location/Qualifiers
     source          1..16657
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(95..683)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(175..194)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(854..1645,1646..1714))
                     /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     1477..1496
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1715..1819)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     1887..1905
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(1943..1965)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2065..2084
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2278..2300
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2292..2309
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2293..2309
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   3013..3075
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     protein_bind    4608..5038
                     /label=gypsy insulator
                     /bound_moiety="Su(Hw), CP190, andMod(mdg4)67.2"
                     /note="chromatin insulator from Drosophila"
     CDS             7277..7342
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             7349..7369
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7394..11494
                     /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             7394..11494
                     /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             11495..11542
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             11543..11596
                     /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(11597..11599,11600..11602,11603..12316)
                     /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(11903..11922)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     12250..12271
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     regulatory      13264..13273
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(13403..13421)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     13671..13690
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     13865..13888
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     polyA_signal    14822..14943
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(14859..14878)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     14913..14932
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     protein_bind    15021..15120
                     /label=phage phi-C31 attP
                     /bound_moiety="phage phi-C31 integrase"
                     /note="attachment site of phage phi-C31"
     primer_bind     complement(16350..16366)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(16350..16366)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(16363..16385)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    16374..16390
                     /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(16398..16404,16405..16422,16423..16428))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    16443..16464
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(16581..16598)
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
       61 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
      121 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
      181 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
      241 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
      301 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
      361 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
      421 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
      481 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
      541 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
      601 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
      661 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
      721 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
      781 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
      841 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
      901 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
      961 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     1021 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     1081 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     1141 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     1201 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     1261 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     1321 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     1381 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     1441 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     1501 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     1561 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     1621 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     1681 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     1741 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     1801 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     1861 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcccttt cgtctcgcgc
     1921 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     1981 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     2041 ggtgtcgggg ctggcttaac tatgcggcat cagagcagat tgtactgaga gtgcaccata
     2101 tgcggtgtga aataccgcac agatgcgtaa ggagaaaata ccgcatcagg cgccattcgc
     2161 cattcaggct gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc
     2221 agctggcgaa agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc
     2281 agtcacgacg ttgtaaaacg acggccagtg ccaagctttg tttaaaaata taacaaaatt
     2341 gtgatcccac aaaatgaagt ggggcaaaat caaataatta atagtgtccg taaacttgtt
     2401 ggtcttcaac tttttgagga acacgttgga cggcaaatcc gtgactataa cacaagttga
     2461 tttaataatt ttagccaaca cgtcgggctg cgtgtttttt gccgacgcgt ctgtgtacac
     2521 gttgattaac tggtcgatta aactgttgaa ataatttaat ttttggttct tctttaaatc
     2581 tgtgatgaaa ttttttaaaa taactttaaa ttcttcattg gtaaaaaatg ccacgttttg
     2641 caacttgtga gggtctaata tgaggtcaaa ctcagtagga gttttatcca aaaaagaaaa
     2701 catgattacg tctgtacacg aacgcgtatt aacgcagagt gcaaagtata agagggttaa
     2761 aaaatatatt ttacgcacca tatacgcatc gggttgatat cgttaatatg gatcaatttg
     2821 aacagttgat taacgtgtct ctgctcaagt ctttgatcaa aacgcaaatc gacgaaaatg
     2881 tgtcggacaa tatcaagtcg atgagcgaaa aactaaaaag gctagaatac gacaatctca
     2941 cagacagcgt tgagatatac ggtattcacg acagcaggct gaataataaa aaaattagaa
     3001 actattattt aaccctagaa agataatcat attgtgacgt acgttaaaga taatcatgcg
     3061 taaaattgac gcatgtgttt tatcggtctg tatatcgagg tttatttatt aatttgaata
     3121 gatattaagt tttattatat ttacacttac atactaataa taaattcaac aaacaattta
     3181 tttatgttta tttatttatt aaaaaaaaac aaaaactcaa aatttcttct ataaagtaac
     3241 aaaactttta aacattctct cttttacaaa aataaactta ttttgtactt taaaaacagt
     3301 catgttgtat tataaaataa gtaattagct taacttatac ataatagaaa caaattatac
     3361 ttattagtca gtcagaaaca actttggcac atatcaatat tatgctctcg acaaataact
     3421 tttttgcatt ttttgcacga tgcatttgcc tttcgcctta ttttagaggg gcagtaagta
     3481 cagtaagtac gttttttcat tactggctct tcagtactgt catctgatgt accaggcact
     3541 tcatttggca aaatattaga gatattatcg cgcaaatatc tcttcaaagt aggagcttct
     3601 aaacgcttac gcataaacga tgacgtcagg ctcatgtaaa ggtttctcat aaattttttg
     3661 cgactttgaa ccttttctcc cttgctactg acattatggc tgtatataat aaaagaattt
     3721 atgcaggcaa tgtttatcat tccgtacaat aatgccatag gccacctatt cgtcttccta
     3781 ctgcaggtgt acctcgccca gttggtccta ttggttgagg gcctacacgg tcgctcgatg
     3841 tgttgtagtt ggtgcaaaag atgaaagatt accggacagt gaccagatga gagttctgga
     3901 agtgcctcag ctgtaatcat tgattgggat atcctttttg tgatggcaaa gtcaatcaaa
     3961 tctggaagct tcatgggatc tgctggccat tatgtaggcg ctcccgggga aacatagtta
     4021 agtttgtgtt gcggtttgat gattgcgttg tacagcaaca aacctttcac caaacgagtg
     4081 aaaagtgttg tgaatttttt tctccaacgc caacgcttaa ctatactcta taactactca
     4141 ttcaccaaaa tctatttgct ctatccataa actagttctt tccaagtaac ttaatctccc
     4201 gaatcgtaat ttctcactcg aactatggcg cgcatagttg tgcttaaggt tggaggtgag
     4261 gtgttttgta gatgcccgaa gtccacctgt ggcttgttcg gtgacaaaaa ctgatctgat
     4321 tctatattct agctctggtc tcaattagac ctccgctcag agaagtaaat aggaaatagt
     4381 gtcttaaatg ctacatttgt aagtgttaat tcataagtgc tgcgtcgcac ttattccgaa
     4441 ttgctgtgtt gcctactctt gagtggatcg attctttgct gagtcgttct cctattacat
     4501 agtccggagg agtaggggtt tcctgtccaa atgtcatgtt aacggccgtt cgctgaacgg
     4561 tcgttgtgtg gcctaattaa atccgttgtt tacattttgt ttaaacttgg ccacgtaata
     4621 agtgtgcgtt gaatttattc gcaaaaacat tgcatatttt cggcaaagta aaattttgtt
     4681 gcatacctta tcaaaaaata agtgctgcat actttttaga gaaaccaaat aattttttat
     4741 tgcatacccg tttttaataa aatacattgc ataccctctt ttaataaaaa atattgcata
     4801 ctttgacgaa acaaattttc gttgcatacc caataaaaga ttattatatt gcatacccgt
     4861 ttttaataaa atacattgca taccctcttt taataaagaa tattgcatac gttgacgaaa
     4921 caaattttcg ttgcataccc aataaaagat tattatattg catacctttt cttgccatac
     4981 catttagccg atcaattgtg ctcggcaaca gtatatttgt ggtgtgccaa ccaacaacgg
     5041 cgcgcctgca gctggttgta ggtgcagttg cgcttcttgc acttggcgca cttgaggaga
     5101 tcggtcttgg tgccctgcac ggtggccagc tgggcatcgt tgatggcctc cttgacgaac
     5161 ttctcgcgca gcttcttcat ctcgtcgctg gccatctcct ccggcgtcat tttggccagc
     5221 tgcttggcag tgacggcgcc gcacataaag ttgccgcgca atccaggatt cttggggtcc
     5281 ttcagattgg ccacgcgtga cctaatgcga ttcttgtact tcatatccgt gttattgaac
     5341 tcggagtaaa tggcatcctc cagttcggcg gccatttcct ccggctcacc gcatccctcg
     5401 ggcacttcgc caatcttcag tgcggtggcc agcatttcgc ggcatttgat gcggaccgca
     5461 tctgtcatgc caccggaggg aaacgaggtc tgtgacgagg atgtcgagcc tttcttctct
     5521 ttatccttgg atgagctgga tgaactggac ttatccttgc cggagattga cgacgacgac
     5581 ttggcggcac tggtggactt cgaggcactg ctgttgttgg aggagccctc cttagcggag
     5641 ctgttgttcg gtgtggttgg cgccgggctg gccaggaagc gtttccagtt cttgatcagc
     5701 gtcttggcca gagcgatcac ctcgtcgtcc ttgctactct tgcgcagctc gtttacggtc
     5761 atgccgatgc gcgttttggt cagaatgtcg agattgatgt taagcgtttg cagggccttc
     5821 agcaggtcca gagcctgatc ctgtccctgt acggtggata tatgtatatt tgtgacatga
     5881 tgcgcggcac gttagctaat cccgatactt actgtgccgt cgctggccat cttgctcatc
     5941 ttcttttgga ttcgaaacac ttcctcctcc acgctcattt tggcccactt aatttgatca
     6001 gaggttaggt taactggcgt cggctgtcgg ctttaatcgg tgctgcgggt ctgcgacagt
     6061 tgtttatgcg gaagccgggc tatttctttt tttccctcca ataaatgttg tttactattt
     6121 ttttcgtttc tgtattcgca gaacaagttg acagaacacc ctaacgcgtt ttagtgttac
     6181 cgggcggtgg cttgtcaaga aataccttcc catggctaat acaattccca actggaagta
     6241 gtttgggttt cagaagtcaa cagtgttacc gtattttaat gtacatctgg cttttaagcg
     6301 agtacacaca catagcattg cgatgttttt taaacttgca ctgcttgcaa gcaggggtcc
     6361 actgtatttc aatgaaagaa cagcgtgttg ggcgtttttg aattatacta attaccttca
     6421 ataacaattc cctatatatc acttagtttt aataaataaa tcgtttgagt gtggtctaga
     6481 gatgatttat tatgatgaag tgcggttgtt ggggacaagc gggatacgtc tgcgggtcaa
     6541 atcggcggtg ggacatagtg agagggcaac ggcgaacaat caccaacgcg ccctggtacg
     6601 cgagaaatgt tgatatcgca catgacctca gcatccacct tcagtttgag aacaaattaa
     6661 atcaggcaca taccagacaa ccaggagaac actgaggagg cgtcatcaaa gatatctttg
     6721 gacacgtggc ataaacaagc caaactatta tctaatcaat atttttatac aaattttgta
     6781 tgtttccatg ttatcttcgt tgctcctgtt agttactacg tttcatagcc cttaaattgg
     6841 ttgcttaacg taaaataaaa ttatattgta taaaaataaa gaggaattca cctataacaa
     6901 acgaaaaagt aaataaataa gaacttcccc atgttaaaaa tgccaccacc atctttacat
     6961 ataagtatgt acatatgaat gaatcactta ggttgcttga atattacaat ttcattaata
     7021 gctaaatcac atttgatgtg ttagtggaaa acggctatat atataaatta tcgaaattgt
     7081 gaatatcgaa ttgcgatagc acaatgggaa attccaccac tagatttttg gtacttttaa
     7141 cagatccttt tcggttttgc gttgcgcgaa gtgatctgaa cttatcaaag tttgtaaggt
     7201 aatacataaa gtgaaaaaga attaatttgc tcttgaaagg caggccaaat taaaaaaaaa
     7261 atatcaattc gagatggact ataaggacca cgacggagac tacaaggatc atgatattga
     7321 ttacaaagac gatgacgata agatggcccc aaagaagaag cggaaggtcg gtatccacgg
     7381 agtcccagca gccgacaaga agtacagcat cggcctggac atcggcacca actctgtggg
     7441 ctgggccgtg atcaccgacg agtacaaggt gcccagcaag aaattcaagg tgctgggcaa
     7501 caccgaccgg cacagcatca agaagaacct gatcggagcc ctgctgttcg acagcggcga
     7561 aacagccgag gccacccggc tgaagagaac cgccagaaga agatacacca gacggaagaa
     7621 ccggatctgc tatctgcaag agatcttcag caacgagatg gccaaggtgg acgacagctt
     7681 cttccacaga ctggaagagt ccttcctggt ggaagaggat aagaagcacg agcggcaccc
     7741 catcttcggc aacatcgtgg acgaggtggc ctaccacgag aagtacccca ccatctacca
     7801 cctgagaaag aaactggtgg acagcaccga caaggccgac ctgcggctga tctatctggc
     7861 cctggcccac atgatcaagt tccggggcca cttcctgatc gagggcgacc tgaaccccga
     7921 caacagcgac gtggacaagc tgttcatcca gctggtgcag acctacaacc agctgttcga
     7981 ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc atcctgtctg ccagactgag
     8041 caagagcaga cggctggaaa atctgatcgc ccagctgccc ggcgagaaga agaatggcct
     8101 gttcggaaac ctgattgccc tgagcctggg cctgaccccc aacttcaaga gcaacttcga
     8161 cctggccgag gatgccaaac tgcagctgag caaggacacc tacgacgacg acctggacaa
     8221 cctgctggcc cagatcggcg accagtacgc cgacctgttt ctggccgcca agaacctgtc
     8281 cgacgccatc ctgctgagcg acatcctgag agtgaacacc gagatcacca aggcccccct
     8341 gagcgcctct atgatcaaga gatacgacga gcaccaccag gacctgaccc tgctgaaagc
     8401 tctcgtgcgg cagcagctgc ctgagaagta caaagagatt ttcttcgacc agagcaagaa
     8461 cggctacgcc ggctacattg acggcggagc cagccaggaa gagttctaca agttcatcaa
     8521 gcccatcctg gaaaagatgg acggcaccga ggaactgctc gtgaagctga acagagagga
     8581 cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc ccccaccaga tccacctggg
     8641 agagctgcac gccattctgc ggcggcagga agatttttac ccattcctga aggacaaccg
     8701 ggaaaagatc gagaagatcc tgaccttccg catcccctac tacgtgggcc ctctggccag
     8761 gggaaacagc agattcgcct ggatgaccag aaagagcgag gaaaccatca ccccctggaa
     8821 cttcgaggaa gtggtggaca agggcgcttc cgcccagagc ttcatcgagc ggatgaccaa
     8881 cttcgataag aacctgccca acgagaaggt gctgcccaag cacagcctgc tgtacgagta
     8941 cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg accgagggaa tgagaaagcc
     9001 cgccttcctg agcggcgagc agaaaaaggc catcgtggac ctgctgttca agaccaaccg
     9061 gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag aaaatcgagt gcttcgactc
     9121 cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc ctgggcacat accacgatct
     9181 gctgaaaatt atcaaggaca aggacttcct ggacaatgag gaaaacgagg acattctgga
     9241 agatatcgtg ctgaccctga cactgtttga ggacagagag atgatcgagg aacggctgaa
     9301 aacctatgcc cacctgttcg acgacaaagt gatgaagcag ctgaagcggc ggagatacac
     9361 cggctggggc aggctgagcc ggaagctgat caacggcatc cgggacaagc agtccggcaa
     9421 gacaatcctg gatttcctga agtccgacgg cttcgccaac agaaacttca tgcagctgat
     9481 ccacgacgac agcctgacct ttaaagagga catccagaaa gcccaggtgt ccggccaggg
     9541 cgatagcctg cacgagcaca ttgccaatct ggccggcagc cccgccatta agaagggcat
     9601 cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg atgggccggc acaagcccga
     9661 gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc cagaagggac agaagaacag
     9721 ccgcgagaga atgaagcgga tcgaagaggg catcaaagag ctgggcagcc agatcctgaa
     9781 agaacacccc gtggaaaaca cccagctgca gaacgagaag ctgtacctgt actacctgca
     9841 gaatgggcgg gatatgtacg tggaccagga actggacatc aaccggctgt ccgactacga
     9901 tgtggaccat atcgtgcctc agagctttct gaaggacgac tccatcgaca acaaggtgct
     9961 gaccagaagc gacaagaacc ggggcaagag cgacaacgtg ccctccgaag aggtcgtgaa
    10021 gaagatgaag aactactggc ggcagctgct gaacgccaag ctgattaccc agagaaagtt
    10081 cgacaatctg accaaggccg agagaggcgg cctgagcgaa ctggataagg ccggcttcat
    10141 caagagacag ctggtggaaa cccggcagat cacaaagcac gtggcacaga tcctggactc
    10201 ccggatgaac actaagtacg acgagaatga caagctgatc cgggaagtga aagtgatcac
    10261 cctgaagtcc aagctggtgt ccgatttccg gaaggatttc cagttttaca aagtgcgcga
    10321 gatcaacaac taccaccacg cccacgacgc ctacctgaac gccgtcgtgg gaaccgccct
    10381 gatcaaaaag taccctaagc tggaaagcga gttcgtgtac ggcgactaca aggtgtacga
    10441 cgtgcggaag atgatcgcca agagcgagca ggaaatcggc aaggctaccg ccaagtactt
    10501 cttctacagc aacatcatga actttttcaa gaccgagatt accctggcca acggcgagat
    10561 ccggaagcgg cctctgatcg agacaaacgg cgaaaccggg gagatcgtgt gggataaggg
    10621 ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc caagtgaata tcgtgaaaaa
    10681 gaccgaggtg cagacaggcg gcttcagcaa agagtctatc ctgcccaaga ggaacagcga
    10741 taagctgatc gccagaaaga aggactggga ccctaagaag tacggcggct tcgacagccc
    10801 caccgtggcc tattctgtgc tggtggtggc caaagtggaa aagggcaagt ccaagaaact
    10861 gaagagtgtg aaagagctgc tggggatcac catcatggaa agaagcagct tcgagaagaa
    10921 tcccatcgac tttctggaag ccaagggcta caaagaagtg aaaaaggacc tgatcatcaa
    10981 gctgcctaag tactccctgt tcgagctgga aaacggccgg aagagaatgc tggcctctgc
    11041 cggcgaactg cagaagggaa acgaactggc cctgccctcc aaatatgtga acttcctgta
    11101 cctggccagc cactatgaga agctgaaggg ctcccccgag gataatgagc agaaacagct
    11161 gtttgtggaa cagcacaagc actacctgga cgagatcatc gagcagatca gcgagttctc
    11221 caagagagtg atcctggccg acgctaatct ggacaaagtg ctgtccgcct acaacaagca
    11281 ccgggataag cccatcagag agcaggccga gaatatcatc cacctgttta ccctgaccaa
    11341 tctgggagcc cctgccgcct tcaagtactt tgacaccacc atcgaccgga agaggtacac
    11401 cagcaccaaa gaggtgctgg acgccaccct gatccaccag agcatcaccg gcctgtacga
    11461 gacacggatc gacctgtctc agctgggagg cgacaaaagg ccggcggcca cgaaaaaggc
    11521 cggccaggca aaaaagaaaa aggagggcag aggaagtctt ctaacatgcg gtgacgtgga
    11581 ggagaatccc ggccctatgg tgagcaaggg cgaggagctg ttcaccgggg tggtgcccat
    11641 cctggtcgag ctggacggcg acgtaaacgg ccacaagttc agcgtgtccg gcgagggcga
    11701 gggcgatgcc acctacggca agctgaccct gaagttcatc tgcaccacca gcaagctgcc
    11761 cgtgccctgg cccaccctcg tgaccaccct gacctacggc gtgcagtgct tcagccgcta
    11821 ccccgaccac atgaagcagc acgacttctt caagtccgcc atgcccgaag gctacgtcca
    11881 ggagcgcacc atcttcttca aggacgacgg caactacaag acccgcgccg aggtgaagtt
    11941 cgagggcgac accctggtga accgcatcga gctgaagggc atcgacttca aggaggacgg
    12001 caacatcctg gggcacaagc tggagtacaa ctacaacagc cacaacgtct atatcatggc
    12061 cgacaagcag aagaacggca tcaaggtgaa cttcaagatc cgccacaaca tcgaggacgg
    12121 cagcgtgcag ctcgccgacc actaccagca gaacaccccc atcggcgacg gccccgtgct
    12181 gctgcccgac aaccactacc tgagcaccca gtccgccctg agcaaagacc ccaacgagaa
    12241 gcgcgatcac atggtcctgc tggagttcgt gaccgccgcc gggatcactc tcggcatgga
    12301 cgagctgtac aagtaattaa ttaactagaa tgaatcgttt ttaaaataac aaatcaattg
    12361 ttttataata ttcgtacgat tctttgatta tgtaataaaa tgtgatcatt aggaagatta
    12421 cgaaaaatat aaaaaatatg agttctgtgt gtataacaaa tgctgtaaac gccacaattg
    12481 tgtttgttgc aaataaaccc atgattattt gattaaaatt gttgttttct ttgttcatag
    12541 acaatagtgt gttttgccta aacgtgtact gcataaactc catgcgagtg tatagcgagc
    12601 tagtggctaa cgcttgcccc accaaagtag attcgtcaaa atcctcaatt tcatcaccct
    12661 cctccaagtt taacatttgg ccgtcggaat taacttctaa agatgccaca taatctaata
    12721 aatgaaatag agattcaaac gtggcgtcat cgtccgtttc gaccatttcc gaaaagaact
    12781 cgggcataaa ctctatgatt tctctggacg tggtgttgtc gaaactctca aagtacgcag
    12841 tcaggaacgt gcgcgacatg tcgtcgggaa actcgcgcgg aaacatgttg ttgtaaccga
    12901 acgggtccca tagcgccaaa accaaatctg ccagcgtcaa tagaatgagc acgatgccga
    12961 caatggagct ggcttggata gcgattcgag ttaacggccg gcagcaattc gagctcgccc
    13021 ggggatctaa ttcaattaga gactaattca attagagcta attcaattag gatccaagct
    13081 tatcgatttc gaaccctcga ccgccggagt ataaatagag gcgcttcgtc tacggagcga
    13141 caattcaatt caaacaagca aagtgaacac gtcgctaagc gaaagctaag caaataaaca
    13201 agcgcagctg aacaagctaa acaatcgggg taccgctaga gtcgacggta cgatccaccg
    13261 gtcgccacca tggtgagcaa gggcgaggag gtcatcaaag agttcatgcg cttcaaggtg
    13321 cgcatggagg gctccatgaa cggccacgag ttcgagatcg agggcgaggg cgagggccgc
    13381 ccctacgagg gcacccagac cgccaagctg aaggtgacca agggcggccc cctgcccttc
    13441 gcctgggaca tcctgtcccc ccagttcatg tacggctcca aggcgtacgt gaagcacccc
    13501 gccgacatcc ccgattacaa gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg
    13561 atgaacttcg aggacggcgg tctggtgacc gtgacccagg actcctccct gcaggacggc
    13621 acgctgatct acaaggtgaa gatgcgcggc accaacttcc cccccgacgg ccccgtaatg
    13681 cagaagaaga ccatgggctg ggaggcctcc accgagcgcc tgtacccccg cgacggcgtg
    13741 ctgaagggcg agatccacca ggccctgaag ctgaaggacg gcggccacta cctggtggag
    13801 ttcaagacca tctacatggc caagaagccc gtgcaactgc ccggctacta ctacgtggac
    13861 accaagctgg acatcacctc ccacaacgag gactacacca tcgtggaaca gtacgagcgc
    13921 tccgagggcc gccaccacct gttcctgggg catggcaccg gcagcaccgg cagcggcagc
    13981 tccggcaccg cctcctccga ggacaacaac atggccgtta tcaaggaatt tatgcgcttc
    14041 aaagttagga tggagggatc catgaacgga catgagttcg agatcgaggg agagggcgag
    14101 ggacgcccgt atgaaggcac acaaacagcc aaactcaagg tcaccaaggg cggaccactg
    14161 cccttcgcct gggatatcct gagtccccag tttatgtacg gcagcaaggc ctacgttaag
    14221 caccccgctg acataccgga ctacaaaaag ctgtcctttc cggaaggctt caagtgggag
    14281 cgcgtgatga atttcgaaga cggaggactg gtcactgtga cccaagatag cagtttgcag
    14341 gacggtacac tgatctataa ggttaaaatg cgcggcacta actttccgcc agatggccca
    14401 gtgatgcaga agaagaccat gggttgggag gcatccaccg aacgtctgta ccctcgagac
    14461 ggagtgctca agggcgagat ccatcaggcc ctcaaactga aagatggtgg tcactacctg
    14521 gtcgaattta agaccattta catggccaag aagccggttc agctgcccgg atattattat
    14581 gtggatacga aactggatat aacttcgcat aacgaagact acaccattgt cgagcagtat
    14641 gagcgcagcg aaggccgaca tcacctgttc ctctacggca tggacgagct gtacaagtag
    14701 gcggccgcga ctctagatca taatcagcca taccacattt gtagaggttt tacttgcttt
    14761 aaaaaacctc ccacacctcc ccctgaacct gaaacataaa atgaatgcaa ttgttgttgt
    14821 taacttgttt attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac
    14881 aaataaagca tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc
    14941 ttaaagcttc ctagggtact gacggacaca ccgaagcccc ggcggcaacc ctcagcggat
    15001 gccccggggc ttcacgtttt cccaggtcag aagcggtttt cgggagtagt gccccaactg
    15061 gggtaacctt tgagttctct cagttggggg cgtagggtcg ccgacatgac acaaggggtt
    15121 gtgaccgggg tggacacgta cgcgggtgct tacgaccgtc agtcgcgcga gcgcgagggc
    15181 ccgggttttt taaatggtac ccttgcagcg ccacctggcc gcgaagagtt ccattctctg
    15241 cttcgaagta cgaagagaga aagagtactt taaatttcca cattcccgcc ttgcagcgcc
    15301 acctggcctt ggtaatgtag aactaggaag gaaagcacca acacaagatg tcgctctccg
    15361 acgatggaca tgtcgcgtaa aaaatgttcg ataactttca atggttcgat tgaacagaca
    15421 ataagtgtat ttaagacacc agttcttata ttcaaaaatc ctaacaactc acaccttgca
    15481 gcgccacctg gccctagaga ccaccaactc tagagagatc tgtaagccgg cgtggcttag
    15541 ctagagggta tgtgtgccgt ttggaaccgg taactcctcc ctccttggat tgaagtgaca
    15601 gccacaacga actagtagtc ttcactaagc tcggctctgg taccgtttta aatgccactc
    15661 ttcttcttct attgatgagg tatattgtcg tcactactcc aactaatagt aatacgtaaa
    15721 gtgctccgaa atttgtagtc gtaaattttt tccaatatat gatggaatta gaagcatggt
    15781 tggatcttat gttcagctta tgcaggttaa tgtcttctgt gttctcggtg aaatatatat
    15841 tttttataat tggggattgt gcggatcttc ttatggaaat tttggtacaa tttgattcaa
    15901 cggtagtgtt ttgtgtgttt aatattaaaa tgtgagctgg ttctacttct tttataattt
    15961 cttgattccc tatgtcttta gttttaaaat tggcagtttc catcttatgt atgttattga
    16021 gacattcagt tttaatatta cctctgattg ccattgcatt gtcgcagata tggtatttat
    16081 atatcttgtt atatttgttg gaaaagtagt aaaaaacatg ccatatagtt aggtatattt
    16141 atatatttat atttatattt catttatggg taatgggaca atgtgtgtga acttttagta
    16201 attttgtttg aatgcaggga ttcttattct aaagtttatc ttaattccaa tcatgaaagt
    16261 atctaattct aacatttgaa atatgtgttt gtctgacatt tcccgtaact ccgggtcgtt
    16321 taattcttgt ctgagaattc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat
    16381 ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc
    16441 taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
    16501 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
    16561 attgggcgct cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg
    16621 cgagcggtat cagctcactc aaaggcggta atacggt
//
