LOCUS       sequence-169910-       14278 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9 with green fluorescent reporter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14278)
  AUTHORS   Herold
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 14278)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: FUCas9GFP
FEATURES             Location/Qualifiers
     source          1..14278
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     173..190
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     LTR             216..396
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     905..921
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     906..922
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    925..1513
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1031..1042
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1440..1460
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(join(1541..2254,2255..2257,2258..2260))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(1544..2254,2255..2257,2258..2260))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1586..1607)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1935..1954
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     CDS             complement(2261..2314)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             complement(2324..2371)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(2372..6472)
                     /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             complement(2372..6472)
                     /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             complement(6497..6517)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(6524..6589)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     regulatory      6589..6598
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        complement(6646..7857)
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     complement(6790..6809)
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     misc_feature    7922..8039
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     CDS             complement(8337..8381)
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    8566..8799
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     misc_feature    9292..9417
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     LTR             9464..9644
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     promoter        complement(9662..9860)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(9687..9707)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        9862..10241
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     10413..10432
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        10507..10611
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10612..10680,10681..11472)
                     /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(10830..10849)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11643..12231
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     12132..12151
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     12385..12402
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    12519..12540
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(12555..12560,12561..12578,12579..12585)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12593..12609
                     /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)."
     primer_bind     12598..12620
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     12617..12633
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     12617..12633
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    12682..12803
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12693..12712)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     12747..12766
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(12933..13307)
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     promoter        complement(13326..13373)
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     promoter        complement(13421..13750)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      13435..13570
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(13489..13508)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     13735..13755
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(13764..14192)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(13874..13895)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     14086..14105
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    join(14238..14278,1..184)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 tcccaatcct cccccttgct gtcctgcccc accccacccc ccagaataga atgacaccta
       61 ctcagacaat gcgatgcaat ttcctcattt tattaggaaa ggacagtggg agtggcacct
      121 tccagggtca aggaaggcac gggggagggg caaacaacag atggctggca actagaaggc
      181 acagtcgagg ctgatcagcg ggtttaaacg ggccctgcta gagattttcc acactgacta
      241 aaagggtctg agggatctct agttaccaga gtcacacaac agacgggcac acactacttg
      301 aagcactcaa ggcaagcttt attgaggctt aagcagtggg ttccctagtt agccagagag
      361 ctcccaggct cagatctggt ctaaccagag agacccagta cagtccggat gcagctctcg
      421 ggccatgtga tgaaatgcta ggcggctgtc aaacctccac tctaatactt ctctctccgg
      481 gtcatccatc ccatgcaggc tcacagggtg taacaagcgg gtgttctctc cttcattggc
      541 ttcttctacc ttctcttgct caactggtac tagcttgtag caccatccaa aggtcagtgg
      601 atatctgatc cctggccctg gtgtgtagtt ctgccaatca gggaagtagc cttgtgtgtg
      661 gtagatccac agatcaagga tatcttgtct tcgttgggag tgaattagcc cttccagtcc
      721 ccccttttct tttaaaaagt ggctaagatc tacagctgcc ttgtaagtca ttggtcttaa
      781 aggtacctga ggtgtgactg gaaaacccac ctcctcctcc tcttgtgctt ctagccaggc
      841 acaatcagca ttggtagctg ctgtattgct acttgtgatt gctccatgtt tttctaggtc
      901 tcgatcgagg tcgacggtat cgatgcgggg aggcggccca aagggagatc cgactcgtct
      961 gagggcgaag gcgaagacgc ggaagaggcc gcagagccgg cagcaggccg cgggaaggaa
     1021 ggtccgctgg attgagggcc gaagggacgt agcagaagga cgtcccgcgc agaatccagg
     1081 tggcaacaca ggcgagcagc caaggaaagg acgatgattt ccccgacaac accacggaat
     1141 tgtcagtgcc caacagccga gcccctgtcc agcagcgggc aaggcaggcg gcgatgagtt
     1201 ccgccgtggc aatagggagg gggaaagcga aagtcccgga aaggagctga caggtggtgg
     1261 caatgcccca accagtgggg gttgcgtcag caaacacagt gcacaccacg ccacgttgcc
     1321 tgacaacggg ccacaactcc tcataaagag acagcaacca ggatttatac aaggaggaga
     1381 aaatgaaagc catacgggaa gcaatagcat gatacaaagg cattaaagca gcgtatccac
     1441 atagcgtaaa aggagcaaca tagttaagaa taccagtcaa tctttcacaa attttgtaat
     1501 ccagaggttg attatcgata agcttgatat cgaattgtca tcacttgtac agctcgtcca
     1561 tgccgagagt gatcccggcg gcggtcacga actccagcag gaccatgtga tcgcgcttct
     1621 cgttggggtc tttgctcagg gcggactggg tgctcaggta gtggttgtcg ggcagcagca
     1681 cggggccgtc gccgatgggg gtgttctgct ggtagtggtc ggcgagctgc acgctgccgt
     1741 cctcgatgtt gtggcggatc ttgaagttca ccttgatgcc gttcttctgc ttgtcggcca
     1801 tgatatagac gttgtggctg ttgtagttgt actccagctt gtgccccagg atgttgccgt
     1861 cctccttgaa gtcgatgccc ttcagctcga tgcggttcac cagggtgtcg ccctcgaact
     1921 tcacctcggc gcgggtcttg tagttgccgt cgtccttgaa gaagatggtg cgctcctgga
     1981 cgtagccttc gggcatggcg gacttgaaga agtcgtgctg cttcatgtgg tcggggtagc
     2041 ggctgaagca ctgcacgccg taggtcaggg tggtcacgag ggtgggccag ggcacgggca
     2101 gcttgccggt ggtgcagatg aacttcaggg tcagcttgcc gtaggtggca tcgccctcgc
     2161 cctcgccgga cacgctgaac ttgtggccgt ttacgtcgcc gtccagctcg accaggatgg
     2221 gcaccacccc ggtgaacagc tcctcgccct tgctcaccat tgggccagga ttctcctcga
     2281 cgtcaccgca tgttagcaga cttcctctgc cctcaccgga tccctttttc ttttttgcct
     2341 ggccggcctt tttcgtggcc gccggccttt tgtcgcctcc cagctgagac aggtcgatcc
     2401 gtgtctcgta caggccggtg atgctctggt ggatcagggt ggcgtccagc acctctttgg
     2461 tgctggtgta cctcttccgg tcgatggtgg tgtcaaagta cttgaaggcg gcaggggctc
     2521 ccagattggt cagggtaaac aggtggatga tattctcggc ctgctctctg atgggcttat
     2581 cccggtgctt gttgtaggcg gacagcactt tgtccagatt agcgtcggcc aggatcactc
     2641 tcttggagaa ctcgctgatc tgctcgatga tctcgtccag gtagtgcttg tgctgttcca
     2701 caaacagctg tttctgctca ttatcctcgg gggagccctt cagcttctca tagtggctgg
     2761 ccaggtacag gaagttcaca tatttggagg gcagggccag ttcgtttccc ttctgcagtt
     2821 cgccggcaga ggccagcatt ctcttccggc cgttttccag ctcgaacagg gagtacttag
     2881 gcagcttgat gatcaggtcc tttttcactt ctttgtagcc cttggcttcc agaaagtcga
     2941 tgggattctt ctcgaagctg cttctttcca tgatggtgat ccccagcagc tctttcacac
     3001 tcttcagttt cttggacttg cccttttcca ctttggccac caccagcaca gaataggcca
     3061 cggtggggct gtcgaagccg ccgtacttct tagggtccca gtccttcttt ctggcgatca
     3121 gcttatcgct gttcctcttg ggcaggatag actctttgct gaagccgcct gtctgcacct
     3181 cggtcttttt cacgatattc acttggggca tgctcagcac tttccgcacg gtggcaaaat
     3241 cccggccctt atcccacacg atctccccgg tttcgccgtt tgtctcgatc agaggccgct
     3301 tccggatctc gccgttggcc agggtaatct cggtcttgaa aaagttcatg atgttgctgt
     3361 agaagaagta cttggcggta gccttgccga tttcctgctc gctcttggcg atcatcttcc
     3421 gcacgtcgta caccttgtag tcgccgtaca cgaactcgct ttccagctta gggtactttt
     3481 tgatcagggc ggttcccacg acggcgttca ggtaggcgtc gtgggcgtgg tggtagttgt
     3541 tgatctcgcg cactttgtaa aactggaaat ccttccggaa atcggacacc agcttggact
     3601 tcagggtgat cactttcact tcccggatca gcttgtcatt ctcgtcgtac ttagtgttca
     3661 tccgggagtc caggatctgt gccacgtgct ttgtgatctg ccgggtttcc accagctgtc
     3721 tcttgatgaa gccggcctta tccagttcgc tcaggccgcc tctctcggcc ttggtcagat
     3781 tgtcgaactt tctctgggta atcagcttgg cgttcagcag ctgccgccag tagttcttca
     3841 tcttcttcac gacctcttcg gagggcacgt tgtcgctctt gccccggttc ttgtcgcttc
     3901 tggtcagcac cttgttgtcg atggagtcgt ccttcagaaa gctctgaggc acgatatggt
     3961 ccacatcgta gtcggacagc cggttgatgt ccagttcctg gtccacgtac atatcccgcc
     4021 cattctgcag gtagtacagg tacagcttct cgttctgcag ctgggtgttt tccacggggt
     4081 gttctttcag gatctggctg cccagctctt tgatgccctc ttcgatccgc ttcattctct
     4141 cgcggctgtt cttctgtccc ttctgggtgg tctggttctc tctggccatt tcgatcacga
     4201 tgttctcggg cttgtgccgg cccatcactt tcacgagctc gtccaccacc ttcactgtct
     4261 gcaggatgcc cttcttaatg gcggggctgc cggccagatt ggcaatgtgc tcgtgcaggc
     4321 tatcgccctg gccggacacc tgggctttct ggatgtcctc tttaaaggtc aggctgtcgt
     4381 cgtggatcag ctgcatgaag tttctgttgg cgaagccgtc ggacttcagg aaatccagga
     4441 ttgtcttgcc ggactgcttg tcccggatgc cgttgatcag cttccggctc agcctgcccc
     4501 agccggtgta tctccgccgc ttcagctgct tcatcacttt gtcgtcgaac aggtgggcat
     4561 aggttttcag ccgttcctcg atcatctctc tgtcctcaaa cagtgtcagg gtcagcacga
     4621 tatcttccag aatgtcctcg ttttcctcat tgtccaggaa gtccttgtcc ttgataattt
     4681 tcagcagatc gtggtatgtg cccagggagg cgttgaaccg atcttccacg ccggagattt
     4741 ccacggagtc gaagcactcg attttcttga agtagtcctc tttcagctgc ttcacggtca
     4801 ctttccggtt ggtcttgaac agcaggtcca cgatggcctt tttctgctcg ccgctcagga
     4861 aggcgggctt tctcattccc tcggtcacgt atttcacttt ggtcagctcg ttatacacgg
     4921 tgaagtactc gtacagcagg ctgtgcttgg gcagcacctt ctcgttgggc aggttcttat
     4981 cgaagttggt catccgctcg atgaagctct gggcggaagc gcccttgtcc accacttcct
     5041 cgaagttcca gggggtgatg gtttcctcgc tctttctggt catccaggcg aatctgctgt
     5101 ttcccctggc cagagggccc acgtagtagg ggatgcggaa ggtcaggatc ttctcgatct
     5161 tttcccggtt gtccttcagg aatgggtaaa aatcttcctg ccgccgcaga atggcgtgca
     5221 gctctcccag gtggatctgg tgggggatgc tgccgttgtc gaaggtccgc tgcttccgca
     5281 gcaggtcctc tctgttcagc ttcacgagca gttcctcggt gccgtccatc ttttccagga
     5341 tgggcttgat gaacttgtag aactcttcct ggctggctcc gccgtcaatg tagccggcgt
     5401 agccgttctt gctctggtcg aagaaaatct ctttgtactt ctcaggcagc tgctgccgca
     5461 cgagagcttt cagcagggtc aggtcctggt ggtgctcgtc gtatctcttg atcatagagg
     5521 cgctcagggg ggccttggtg atctcggtgt tcactctcag gatgtcgctc agcaggatgg
     5581 cgtcggacag gttcttggcg gccagaaaca ggtcggcgta ctggtcgccg atctgggcca
     5641 gcaggttgtc caggtcgtcg tcgtaggtgt ccttgctcag ctgcagtttg gcatcctcgg
     5701 ccaggtcgaa gttgctcttg aagttggggg tcaggcccag gctcagggca atcaggtttc
     5761 cgaacaggcc attcttcttc tcgccgggca gctgggcgat cagattttcc agccgtctgc
     5821 tcttgctcag tctggcagac aggatggcct tggcgtccac gccgctggcg ttgatggggt
     5881 tttcctcgaa cagctggttg taggtctgca ccagctggat gaacagcttg tccacgtcgc
     5941 tgttgtcggg gttcaggtcg ccctcgatca ggaagtggcc ccggaacttg atcatgtggg
     6001 ccagggccag atagatcagc cgcaggtcgg ccttgtcggt gctgtccacc agtttctttc
     6061 tcaggtggta gatggtgggg tacttctcgt ggtaggccac ctcgtccacg atgttgccga
     6121 agatggggtg ccgctcgtgc ttcttatcct cttccaccag gaaggactct tccagtctgt
     6181 ggaagaagct gtcgtccacc ttggccatct cgttgctgaa gatctcttgc agatagcaga
     6241 tccggttctt ccgtctggtg tatcttcttc tggcggttct cttcagccgg gtggcctcgg
     6301 ctgtttcgcc gctgtcgaac agcagggctc cgatcaggtt cttcttgatg ctgtgccggt
     6361 cggtgttgcc cagcaccttg aatttcttgc tgggcacctt gtactcgtcg gtgatcacgg
     6421 cccagcccac agagttggtg ccgatgtcca ggccgatgct gtacttcttg tcggctgctg
     6481 ggactccgtg gataccgacc ttccgcttct tctttggggc catcttatcg tcatcgtctt
     6541 tgtaatcaat atcatgatcc ttgtagtctc cgtcgtggtc cttatagtcc atggtggcac
     6601 cgtcagaatt cagatcctct agagtcgacc tgcagcccaa gcttcgtcta acaaaaaagc
     6661 caaaaacggc cagaatttag cggacaattt actagtctaa cactgaaaat tacatattga
     6721 cccaaatgat tacatttcaa aaggtgccta aaaaacttca caaaacacac tcgccaaccc
     6781 cgagcgcata gttcaaaacc ggagcttcag ctacttaaga agataggtac ataaaaccga
     6841 ccaaagaaac tgacgcctca cttatccctc ccctcaccag aggtccggcg cctgtcgatt
     6901 caggagagcc taccctaggc ccgaaccctg cgtcctgcga cggagaaaag cctaccgcac
     6961 acctaccggc aggtggcccc accctgcatt ataagccaac agaacgggtg acgtcacgac
     7021 acgacgaggg cgcgcgctcc caaaggtacg ggtgcactgc ccaacggcac cgccataact
     7081 gccgcccccg caacagacga caaaccgagt tctccagtca gtgacaaact tcacgtcagg
     7141 gtccccagat ggtgccccag cccatctcac ccgaataaga gctttcccgc attagcgaag
     7201 gcctcaagac cttgggttct tgccgcccac catgcccccc accttgtttc aacgacctca
     7261 cagcccgcct cacaagcgtc ttccattcaa gactcgggaa cagccgccat tttgctgcgc
     7321 tccccccaac ccccagttca gggcaacctt gctcgcggac ccagactaca gcccttggcg
     7381 gtctctccac acgcttccgt cccaccgagc ggcccggcgg ccacgaaagc cccggccagc
     7441 ccagcagccc gctactcacc aagtgacgat cacagcgatc cacaaacaag aaccgcgacc
     7501 caaatcccgg ctgcgacgga actagctgtg ccacacccgg cgcgtcctta tataatcatc
     7561 ggcgttcacc gccccacgga gatccctccg cagaatcgcc gagaagggac tacttttcct
     7621 cgcctgttcc gctctctgga aagaaaacca gtgccctaga gtcacccaag tcccgtccta
     7681 aaatgtcctt ctgctgatac tggggttcta aggccgagtc ttatgagcag cgggccgctg
     7741 tcctgagcgt ccgggcggaa ggatcaggac gctcgctgcg cccttcgtct gacgtggcag
     7801 cgctcgccgt gaggaggggg gcgcccgcgg gaggcgccaa aacccggcgc ggaggccaga
     7861 tctggagccg acacgggtta attaaccaaa ctggatctct gctgtccctg taataaaccc
     7921 gaaaattttg aatttttgta atttgttttt gtaattcttt agtttgtatg tctgttgcta
     7981 ttatgtctac tattctttcc cctgcactgt accccccaat cccccctttt cttttaaaat
     8041 tgtggatgaa tactgccatt tgtctgcaga attggcgcac gcagtgccga tccgttcact
     8101 aatcgaatgg atctgtctct gtctctctct ccaccttctt cttctattcc ttcgggcctg
     8161 tcgggtcccc tcggggttgg gaggtgggtc tgaaacgata atggtgaata tccctgccta
     8221 actctattca ctatagaaag tacagcaaaa actattctta aacctaccaa gcctcctact
     8281 atcattatga ataattttat ataccacagc caatttgtta tgttaaacca attccacaaa
     8341 cttgcccatt tatctaattc caataattct tgttcattct tttcttgctg gttttgcgat
     8401 tcttcaatta aggagtgtat taagcttgtg taattgttaa tttctctgtc ccactccatc
     8461 caggtcgtgt gattccaaat ctgttccaga gatttattac tccaactagc attccaaggc
     8521 acagcagtgg tgcaaatgag ttttccagag caaccccaaa tccccaggag ctgttgatcc
     8581 tttaggtatc tttccacagc caggattctt gcctggagct gcttgatgcc ccagactgtg
     8641 agttgcaaca gatgctgttg cgcctcaata gccctcagca aattgttctg ctgctgcact
     8701 ataccagaca ataattgtct ggcctgtacc gtcagcgtca ttgacgctgc gcccatagtg
     8761 cttcctgctg ctcccaagaa cccaaggaac aaagctccta ttcccactgc tcttttttct
     8821 ctctgcacca ctcttctctt tgccttggtg ggtgctactc ctaatggttc aatttttact
     8881 actttatatt tatataattc acttctccaa ttgtccctca tatctcctcc tccaggtctg
     8941 aagatcagcg gccgcttgct gtgcggtggt cttacttttg ttttgctctt cctctatctt
     9001 gtctaaagct tccttggtgt cttttatctc tatcctttga tgcacacaat agagggttgc
     9061 tactgtatta tataatgatc taagttcttc tgatcctgtc tgaagggatg gttgtagctg
     9121 tcccagtatt tgtctacagc cttctgatgt ttctaacagg ccaggattaa ctgcgaatcg
     9181 ttctagctcc ctgcttgccc atactatatg ttttaattta tattttttct ttccccctgg
     9241 ccttaaccga attttttccc atcgcgatct aattctcccc cgcttaatac tgacgctctc
     9301 gcacccatct ctctccttct agcctccgct agtcaaaatt tttggcgtac tcaccagtcg
     9361 ccgcccctcg cctcttgccg tgcgcgcttc agcaagccga gtcctgcgtc gagagagctc
     9421 ctctggtttc cctttcgctt tcaagtccct gttcgggcgc cactgctaga gattttccac
     9481 actgactaaa agggtctgag ggatctctag ttaccagagt cacacaacag acgggcacac
     9541 actacttgaa gcactcaagg caagctttat tgaggcttaa gcagtgggtt ccctagttag
     9601 ccagagagct cccaggctca gatctggtct aaccagagag acccagtaca ggcaaaacgc
     9661 gctgcttata tagacctccc accgtacacg cctaccgccc atttgcgtca atggggcgga
     9721 gttgttacga cattttggaa agtcccgttg attttggtgc caaaacaaac tcccattgac
     9781 gtcaatgggg tggagacttg gaaatccccg tgagtcaaac cgctatccac gcccattgat
     9841 gtactgccaa aaccgcatca ccatggtaat agcgatgact aatacgtaga tgtactgcca
     9901 agtaggaaag tcccataagg tcatgtactg ggcataatgc caggcgggcc atttaccgtc
     9961 attgacgtca atagggggcg tacttggcat atgatacact tgatgtactg ccaagtgggc
    10021 agtttaccgt aaatactcca cccattgacg tcaatggaaa gtccctattg gcgttactat
    10081 gggaacatac gtcattattg acgtcaatgg gcgggggtcg ttgggcggtc agccaggcgg
    10141 gccatttacc gtaagttatg taacgcggaa ctccatatat gggctatgaa ctaatgaccc
    10201 cgtaattgat tactattaat aactagtcaa taatcaatgt caacgcgtat atctggcccg
    10261 tacatcgcga agcagcgcaa aacgcctaac cctaagcaga ttcttcatgc aattgtcggt
    10321 caagccttgc cttgttgtag cttaaatttt gctcgcgcac tactcagcga cctccaacac
    10381 acaagcaggg agcagatact ggcttaacta tgcggcatca gagcagattg tactgagagt
    10441 gcaccatagg ggatcgggag atctcccgat ccgtcgacgt caggtggcac ttttcgggga
    10501 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
    10561 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
    10621 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
    10681 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
    10741 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
    10801 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
    10861 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
    10921 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
    10981 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
    11041 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
    11101 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
    11161 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
    11221 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
    11281 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
    11341 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
    11401 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
    11461 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
    11521 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
    11581 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
    11641 tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta
    11701 ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc
    11761 ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt aggccaccac
    11821 ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtggct
    11881 gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata gttaccggat
    11941 aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt ggagcgaacg
    12001 acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac gcttcccgaa
    12061 gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg
    12121 gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg ccacctctga
    12181 cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa aaacgccagc
    12241 aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat gttctttcct
    12301 gcgttatccc ctgattctgt ggataaccgt attaccgcct ttgagtgagc tgataccgct
    12361 cgccgcagcc gaacgaccga gcgcagcgag tcagtgagcg aggaagcgga agagcgccca
    12421 atacgcaaac cgcctctccc cgcgcgttgg ccgattcatt aatgcagctg gcacgacagg
    12481 tttcccgact ggaaagcggg cagtgagcgc aacgcaatta atgtgagtta gctcactcat
    12541 taggcacccc aggctttaca ctttatgctt ccggctcgta tgttgtgtgg aattgtgagc
    12601 ggataacaat ttcacacagg aaacagctat gaccatgatt acgccaagct ctagctagag
    12661 gtcgacggta tacagacatg ataagataca ttgatgagtt tggacaaacc acaactagaa
    12721 tgcagtgaaa aaaatgcttt atttgtgaaa tttgtgatgc tattgcttta tttgtaacca
    12781 ttataagctg caataaacaa gttggggtgg gcgaagaact ccagcatgag atccccgcgc
    12841 tggaggatca tccagccggc gtcccggaaa acgattccga agcccaacct ttcatagaag
    12901 gcggcggtgg aatcgaaatc tcgtagcacg tgtcagtcct gctcctcggc cacgaagtgc
    12961 acgcagttgc cggccgggtc gcgcagggcg aactcccgcc cccacggctg ctcgccgatc
    13021 tcggtcatgg ccggcccgga ggcgtcccgg aagttcgtgg acacgacctc cgaccactcg
    13081 gcgtacagct cgtccaggcc gcgcacccac acccaggcca gggtgttgtc cggcaccacc
    13141 tggtcctgga ccgcgctgat gaacagggtc acgtcgtccc ggaccacacc ggcgaagtcg
    13201 tcctccacga agtcccggga gaacccgagc cggtcggtcc agaactcgac cgctccggcg
    13261 acgtcgcgcg cggtgagcac cggaacggca ctggtcaact tggccatggt ttagttcctc
    13321 accttgtcgt attatactat gccgatatac tatgccgatg attaattgtc aacacgtgct
    13381 gatcagatcc gaaaatggat atacaagctc ccgggagctt tttgcaaaag cctaggcctc
    13441 caaaaaagcc tcctcactac ttctggaata gctcagaggc agaggcggcc tcggcctctg
    13501 cataaataaa aaaaattagt cagccatggg gcggagaatg ggcggaactg ggcggagtta
    13561 ggggcgggat gggcggagtt aggggcggga ctatggttgc tgactaattg agatgcatgc
    13621 tttgcatact tctgcctgct ggggagcctg gggactttcc acacctggtt gctgactaat
    13681 tgagatgcat gctttgcata cttctgcctg ctggggagcc tggggacttt ccacacccta
    13741 actgacacac attccacaga attaattcgc gttaaatttt tgttaaatca gctcattttt
    13801 taaccaatag gccgaaatcg gcaaaatccc ttataaatca aaagaataga ccgagatagg
    13861 gttgagtgtt gttccagttt ggaacaagag tccactatta aagaacgtgg actccaacgt
    13921 caaagggcga aaaaccgtct atcagggcga tggcccacta cgtgaaccat caccctaatc
    13981 aagttttttg gggtcgaggt gccgtaaagc actaaatcgg aaccctaaag ggagcccccg
    14041 atttagagct tgacggggaa agccggcgaa cgtggcgaga aaggaaggga agaaagcgaa
    14101 aggagcgggc gctagggcgc tggcaagtgt agcggtcacg ctgcgcgtaa ccaccacacc
    14161 cgccgcgctt aatgcgccgc tacagggcgc gtggggatac cccctagagc cccagctggt
    14221 tctttccgcc tcagaagcca tagagcccac cgcatcccca gcatgcctgc tattgtct
//
