LOCUS       sequence-453885-       10680 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  miniEDV production plasmid. Expresses the miniGag-Cas9 polypeptide 
            (CAG promoter) and sgRNA (human U6 promoter).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10680)
  AUTHORS   Ngo W, Peukes J, Baldwin A, Xue ZW, Hwang S, Stickels RR, Lin Z, 
            Satpathy AT, Wells JA, Schekman R, Nogales E, Doudna JA
  TITLE     Mechanism-guided engineering of a minimal biological particle for 
            genome editing.
  JOURNAL   Proc Natl Acad Sci U S A. 2025 Jan 7;122(1):e2413519121. doi: 
            10.1073/pnas.2413519121. Epub 2024 Dec 30.
  PUBMED    39793042
REFERENCE   2  (bases 1 to 10680)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2025-01-7"; volume: "122(1):e2413519121. 
            doi"; pages: " 10.1073/pnas.2413519121. Epub 2024 Dec 30"
COMMENT     Sequence Label: pWN_U6-B2M-miniGag-Cas9
FEATURES             Location/Qualifiers
     source          1..10680
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             45..65
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             72..92
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             99..119
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             126..146
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(159..178)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    224..279
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(278..297)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(640..656)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(640..656)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(653..675)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    664..680
                     /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(688..694,695..712,713..718))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    733..754
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        813..1008
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      859..994
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    1014..1148
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1051..1070)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1105..1124
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1216..1233)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1387..1975)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1467..1486)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2146..2937,2938..3006))
                     /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     2769..2788
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3007..3111)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3193..3441
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     3193..3213
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     3364..3383
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     enhancer        3595..3974
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3976..4253
                     /label=chicken beta-actin promoter
     intron          4254..5270
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     5194..5216
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     5278..5297
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      5325..5334
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             6258..6284
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             6297..6323
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             6336..6362
                     /codon_start=1
                     /product="nuclear export signal from the HIV Rev protein 
                     (Fischer et al., 1995)"
                     /label=NES
                     /translation="LPPLERLTL"
     CDS             6534..10637
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             6534..10637
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             join(10641..10680,1..26)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
ORIGIN
        1 ttgattacaa agacgatgac gataagagcc tgggcagcgg ctcccccaag aaaaagagaa
       61 aagtcgaaga ccccaagaag aaaaggaagg tcgaagatcc taaaaagaaa aggaaagtgg
      121 aagatcctaa gaaaaagcgg aaagtgtaag ctagcagatc tttttccctc tgccaaaaat
      181 tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag gaaatttatt
      241 ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac atatgggagg
      301 gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac atatgcccat
      361 atgctggctg ccatgaacaa aggttggcta taaagaggtc atcagtatat gaaacagccc
      421 cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat tttttttata
      481 ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca tgttttacta
      541 gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct tctcttatgg
      601 agatccctcg acctgcagcc caagcttggc gtaatcatgg tcatagctgt ttcctgtgtg
      661 aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa agtgtaaagc
      721 ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac tgcccgcttt
      781 ccagtcggga aacctgtcgt gccagcggat ccgcatctca attagtcagc aaccatagtc
      841 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
      901 catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc ctctgagcta
      961 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctaacttgt
     1021 ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc acaaataaag
     1081 catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta tcttatcatg
     1141 tctggatccg ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg
     1201 gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc
     1261 ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg ataacgcagg
     1321 aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg ccgcgttgct
     1381 ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac gctcaagtca
     1441 gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gaagctccct
     1501 cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct ttctcccttc
     1561 gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg tgtaggtcgt
     1621 tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct gcgccttatc
     1681 cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac tggcagcagc
     1741 cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt tcttgaagtg
     1801 gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc tgctgaagcc
     1861 agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca ccgctggtag
     1921 cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat ctcaagaaga
     1981 tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac gttaagggat
     2041 tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt aaaaatgaag
     2101 ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc aatgcttaat
     2161 cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg cctgactccc
     2221 cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg ctgcaatgat
     2281 accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc cagccggaag
     2341 ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta ttaattgttg
     2401 ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg ttgccattgc
     2461 tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct ccggttccca
     2521 acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta gctccttcgg
     2581 tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg ttatggcagc
     2641 actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga ctggtgagta
     2701 ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt gcccggcgtc
     2761 aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca ttggaaaacg
     2821 ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt cgatgtaacc
     2881 cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt ctgggtgagc
     2941 aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga aatgttgaat
     3001 actcatactc ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag
     3061 cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc gcacatttcc
     3121 ccgaaaagtg ccacctggtc gacgggttta ttacagggac agcagagatc cagtttggtt
     3181 aattaaggta ccgagggcct atttcccatg attccttcat atttgcatat acgatacaag
     3241 gctgttagag agataattag aattaatttg actgtaaaca caaagatatt agtacaaaat
     3301 acgtgacgta gaaagtaata atttcttggg tagtttgcag ttttaaaatt atgttttaaa
     3361 atggactatc atatgcttac cgtaacttga aagtatttcg atttcttggc tttatatatc
     3421 ttgtggaaag gacgaaacac cggagtagcg cgagcacagc tagtttaaga gctatgctgg
     3481 aaacagcata gcaagtttaa ataaggctag tccgttatca acttgaaaaa gtggcaccga
     3541 gtcggtgctt tttttgaatt cgctagctag gtcttgaaag gagtgggaat tgtcgacatt
     3601 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
     3661 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
     3721 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
     3781 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
     3841 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
     3901 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
     3961 tcgctattac catggtcgag gtgagcccca cgttctgctt cactctcccc atctcccccc
     4021 cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca gcgatggggg
     4081 cggggggggg gggggggcgc gcgccaggcg gggcggggcg gggcgagggg cggggcgggg
     4141 cgaggcggag aggtgcggcg gcagccaatc agagcggcgc gctccgaaag tttcctttta
     4201 tggcgaggcg gcggcggcgg cggccctata aaaagcgaag cgcgcggcgg gcgggagtcg
     4261 ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc ctcgcgccgc ccgccccggc
     4321 tctgactgac cgcgttactc ccacaggtga gcgggcggga cggcccttct cctccgggct
     4381 gtaattagcg cttggtttaa tgacggcttg tttcttttct gtggctgcgt gaaagccttg
     4441 aggggctccg ggagggccct ttgtgcgggg ggagcggctc ggggggtgcg tgcgtgtgtg
     4501 tgtgcgtggg gagcgccgcg tgcggctccg cgctgcccgg cggctgtgag cgctgcgggc
     4561 gcggcgcggg gctttgtgcg ctccgcagtg tgcgcgaggg gagcgcggcc gggggcggtg
     4621 ccccgcggtg cggggggggc tgcgagggga acaaaggctg cgtgcggggt gtgtgcgtgg
     4681 gggggtgagc agggggtgtg ggcgcgtcgg tcgggctgca accccccctg cacccccctc
     4741 cccgagttgc tgagcacggc ccggcttcgg gtgcggggct ccgtacgggg cgtggcgcgg
     4801 ggctcgccgt gccgggcggg gggtggcggc aggtgggggt gccgggcggg gcggggccgc
     4861 ctcgggccgg ggagggctcg ggggaggggc gcggcggccc ccggagcgcc ggcggctgtc
     4921 gaggcgcggc gagccgcagc cattgccttt tatggtaatc gtgcgagagg gcgcagggac
     4981 ttcctttgtc ccaaatctgt gcggagccga aatctgggag gcgccgccgc accccctcta
     5041 gcgggcgcgg ggcgaagcgg tgcggcgccg gcaggaagga aatgggcggg gagggccttc
     5101 gtgcgtcgcc gcgccgccgt ccccttctcc ctctccagcc tcggggctgt ccgcgggggg
     5161 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     5221 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     5281 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attcgccacc atgggtgcga
     5341 gagcgtcagt attaagcggg ggagaattag atcgatggga aaaaattcgg ttaaggccag
     5401 ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat
     5461 tcgcagttaa tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc
     5521 tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa
     5581 ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga
     5641 tagaggaaga gcaaaacaaa agtaagaaaa aagcacagca agcagcagct gacacaggac
     5701 acagcaatca ggtcagccaa aattacccta tagtgcagag cattctggac ataagacaag
     5761 gaccaaagga accctttaga gactatgtag accgattcta taaaactcta agagccgagc
     5821 aagcttcaca agaggtaaaa aattggatga cagaaacctt gttggtccaa aatgcgaacc
     5881 cagattgtaa gactatttta aaagcattgg gaccaggagc gacactagaa gaaatgatga
     5941 cagcatgtca gggagtgggg ggacccggcc ataaagcaag agttttggct gaagcaatga
     6001 gccaagtaac aaatccagct accataatga tacagaaagg cagacaggct aattttttag
     6061 ggaagatctg gccttcccac aagggaaggc cagggaattt tcttcagagc agaccagagc
     6121 caacagcccc accagaagag agcttcaggt ttggggaaga gacaacaact ccctctcaga
     6181 agcaggagcc gatagacaag gaactgtatc ctttagcttc cctcagatca ctctttggca
     6241 gcgacccctc gtcacaactc ccccctctgg agcgccttac tcttggctcc ctgcagttgc
     6301 ccccactgga gcggctgacc ctggggtcat tgcagttgcc tccactcgaa cgactgacgt
     6361 tgagccagaa ctatccgatt gtgcagaccg gtatgaagcg agcactgccc aacaacacca
     6421 gctcctctcc ccagccaaag aagaaaccag gagggtcagg gggaaagcga gcactgccca
     6481 acaacaccag ctcctctccc cagccaaaga agaaaccagg agggtcaggg ggaatggaca
     6541 agaagtacag catcggcctg gacatcggca ccaactctgt gggctgggcc gtgatcaccg
     6601 acgagtacaa ggtgcccagc aagaaattca aggtgctggg caacaccgac cggcacagca
     6661 tcaagaagaa cctgatcggc gccctgctgt tcgacagcgg agaaacagcc gaggccaccc
     6721 ggctgaagag aaccgccaga agaagataca ccagacggaa gaaccggatc tgctatctgc
     6781 aagagatctt cagcaacgag atggccaagg tggacgacag cttcttccac agactggaag
     6841 agtccttcct ggtggaagag gataagaagc acgagcggca ccccatcttc ggcaacatcg
     6901 tggacgaggt ggcctaccac gagaagtacc ccaccatcta ccacctgaga aagaaactgg
     6961 tggacagcac cgacaaggcc gacctgcggc tgatctatct ggccctggcc cacatgatca
     7021 agttccgggg ccacttcctg atcgagggcg acctgaaccc cgacaacagc gacgtggaca
     7081 agctgttcat ccagctggtg cagacctaca accagctgtt cgaggaaaac cccatcaacg
     7141 ccagcggcgt ggacgccaag gccatcctgt ctgccagact gagcaagagc agacggctgg
     7201 aaaatctgat cgcccagctg cccggcgaga agaagaatgg cctgttcggc aacctgattg
     7261 ccctgagcct gggcctgacc cccaacttca agagcaactt cgacctggcc gaggatgcca
     7321 aactgcagct gagcaaggac acctacgacg acgacctgga caacctgctg gcccagatcg
     7381 gcgaccagta cgccgacctg tttctggccg ccaagaacct gtccgacgcc atcctgctga
     7441 gcgacatcct gagagtgaac accgagatca ccaaggcccc cctgagcgcc tctatgatca
     7501 agagatacga cgagcaccac caggacctga ccctgctgaa agctctcgtg cggcagcagc
     7561 tgcctgagaa gtacaaagag attttcttcg accagagcaa gaacggctac gccggctaca
     7621 tcgatggcgg agccagccag gaagagttct acaagttcat caagcccatc ctggaaaaga
     7681 tggacggcac cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc
     7741 ggaccttcga caacggcagc atcccccacc agatccacct gggagagctg cacgccattc
     7801 tgcggcggca ggaagatttt tacccattcc tgaaggacaa ccgggaaaag atcgagaaga
     7861 tcctgacctt ccgcatcccc tactacgtgg gccctctggc caggggaaac agcagattcg
     7921 cctggatgac cagaaagagc gaggaaacca tcaccccctg gaacttcgag gaagtggtgg
     7981 acaagggcgc cagcgcccag agcttcatcg agcggatgac caacttcgat aagaacctgc
     8041 ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga gtacttcacc gtgtacaacg
     8101 agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg
     8161 agcagaaaaa agccatcgtg gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc
     8221 agctgaaaga ggactacttc aagaaaatcg agtgcttcga ctccgtggaa atctccggcg
     8281 tggaagatcg gttcaacgcc tccctgggca cataccacga tctgctgaaa attatcaagg
     8341 acaaggactt cctggacaat gaggaaaacg aggacattct ggaagatatc gtgctgaccc
     8401 tgacactgtt tgaggacaga gagatgatcg aggaacggct gaaaacctat gcccacctgt
     8461 tcgacgacaa agtgatgaag cagctgaagc ggcggagata caccggctgg ggcaggctga
     8521 gccggaagct gatcaacggc atccgggaca agcagtccgg caagacaatc ctggatttcc
     8581 tgaagtccga cggcttcgcc aacagaaact tcatgcagct gatccacgac gacagcctga
     8641 cctttaaaga ggacatccag aaagcccagg tgtccggcca gggcgatagc ctgcacgagc
     8701 acattgccaa tctggccggc agccccgcca ttaagaaggg catcctgcag acagtgaagg
     8761 tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa
     8821 tggccagaga gaaccagacc acccagaagg gacagaagaa cagccgcgag agaatgaagc
     8881 ggatcgaaga gggcatcaaa gagctgggca gccagatcct gaaagaacac cccgtggaaa
     8941 acacccagct gcagaacgag aagctgtacc tgtactacct gcagaatggg cgggatatgt
     9001 acgtggacca ggaactggac atcaaccggc tgtccgacta cgatgtggac catatcgtgc
     9061 ctcagagctt tctgaaggac gactccatcg ataacaaagt gctgactcgg agcgacaaga
     9121 accggggcaa gagcgacaac gtgccctccg aagaggtcgt gaagaagatg aagaactact
     9181 ggcgccagct gctgaatgcc aagctgatta cccagaggaa gttcgacaat ctgaccaagg
     9241 ccgagagagg cggcctgagc gaactggata aggccggctt catcaagaga cagctggtgg
     9301 aaacccggca gatcacaaag cacgtggcac agatcctgga ctcccggatg aacactaagt
     9361 acgacgagaa cgacaaactg atccgggaag tgaaagtgat caccctgaag tccaagctgg
     9421 tgtccgattt ccggaaggat ttccagtttt acaaagtgcg cgagatcaac aactaccacc
     9481 acgcccacga cgcctacctg aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta
     9541 agctggaaag cgagttcgtg tacggcgact acaaggtgta cgacgtgcgg aagatgatcg
     9601 ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta cttcttctac agcaacatca
     9661 tgaacttttt caagaccgag attaccctgg ccaacggcga gatccggaag cggcctctga
     9721 tcgagacaaa cggcgaaaca ggcgagatcg tgtgggataa gggccgggac tttgccaccg
     9781 tgcggaaagt gctgtctatg ccccaagtga atatcgtgaa aaagaccgag gtgcagacag
     9841 gcggcttcag caaagagtct atcctgccca agaggaacag cgacaagctg atcgccagaa
     9901 agaaggactg ggaccctaag aagtacggcg gcttcgacag ccccaccgtg gcctattctg
     9961 tgctggtggt ggccaaagtg gaaaagggca agtccaagaa actgaagagt gtgaaagagc
    10021 tgctggggat caccatcatg gaaagaagca gcttcgagaa gaatcccatc gactttctgg
    10081 aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat caagctgcct aagtactccc
    10141 tgttcgagct ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg
    10201 gaaacgaact ggccctgccc tccaaatatg tgaacttcct gtacctggcc agccactatg
    10261 agaagctgaa gggctccccc gaggataatg agcagaaaca gctgtttgtg gaacagcaca
    10321 aacactacct ggacgagatc atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg
    10381 ccgacgctaa tctggacaag gtgctgagcg cctacaacaa gcacagagac aagcctatca
    10441 gagagcaggc cgagaatatc atccacctgt ttaccctgac caatctggga gcccctgccg
    10501 ccttcaagta ctttgacacc accatcgacc ggaagaggta caccagcacc aaagaggtgc
    10561 tggacgccac cctgatccac cagagcatca ccggcctgta cgagacacgg atcgacctgt
    10621 ctcagctggg aggcgacgcc gactataagg accacgacgg agactacaag gatcatgata
//
