LOCUS       sequence-371825-       13168 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PiggyBac compatible plasmid expressing dCas9-KRAB-MeCP2 expression 
            plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13168)
  AUTHORS   Nghi Dang , Alissa Lance-Byrne, Angela Tung, Xiaoge Guo, Ryan J 
            Cecchi, Joanna Buchthal, Alejandro Chavez, Nan Cher Yeo
  TITLE     Generation and application of a versatile CRISPR toolkit for 
            mammalian cell engineering
  JOURNAL   Synthetic Biology, Volume 6, Issue 1, 2021
REFERENCE   2  (bases 1 to 13168)
  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: "Synthetic 
            Biology, Volume 6, Issue 1, 2021"
COMMENT     Sequence Label: NAX_CAGGS_CAS9_m4_KRAB-MeCP2_mcherry
FEATURES             Location/Qualifiers
     source          1..13168
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          1..1017
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     941..963
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1025..1044
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1114..1123
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1120..1146,1147..1149,1150..3633,3634..3639,3640..3705,
                     3706..3708,3709..5223)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=Cas9m4
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the mutations D10A, D839A, 
                     H840A, and N863A (Mali et al., 2013)"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVAAIVPQSFLKDDSIDNKVLTRSDKARGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5236..5256
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5317..5511
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEPWLV"
     CDS             5545..5565
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5575..6432
                     /codon_start=1
                     /product="transcriptional repression domain from rat 
                     methyl-CpG-binding protein 2 (Nan et al., 1997)"
                     /label=MeCP2
                     /translation="VQVKRVLEKSPGKLLVKMPFQASPGGKGEGGGATTSAQVMVIKRP
                     GRKRKAEADPQAIPKKRGRKPGSVVAAAAAEAKKKAVKESSIRSVQETVLPIKKRKTRE
                     TVSIEVKEVVKPLLVSTLGEKSGKGLKTCKSPGRKSKESSPKGRSSSASSPPKKEHHHH
                     HHHAESPKAPMPLLPPPPPPEPQSSEDPISPPEPQDLSSSICKEEKMPRAGSLESDGCP
                     KEPAKTQPMVAAAATTTTTTTTTVAEKYKHRGEGERKDIVSSSMPRPNREEPVDSRTPV
                     TERVS"
     primer_bind     complement(6487..6506)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    6531..6579
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        6811..7022
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      7032..7041
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(7156..7174)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     7424..7443
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    7725..7846
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7762..7781)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7816..7835
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     repeat_region   8609..8643
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(9134..9150)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9134..9150)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(9147..9169)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    9158..9174
                     /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(9182..9188,9189..9206,9207..9212))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9227..9248
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(9365..9382)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(9536..10124)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9616..9635)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(10295..11086,11087..11155))
                     /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     10918..10937
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(11156..11260)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(11286..11741)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11423..11444)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     11635..11654
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11868..11890
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     11882..11899
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     11883..11899
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     repeat_region   12029..12091
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     enhancer        12512..12891
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 ggagtcgctg cgcgctgcct tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg
       61 ccccggctct gactgaccgc gttactccca caggtgagcg ggcgggacgg cccttctcct
      121 ccgggctgta attagcgctt ggtttaatga cggcttgttt cttttctgtg gctgcgtgaa
      181 agccttgagg ggctccggga gggccctttg tgcgggggga gcggctcggg gggtgcgtgc
      241 gtgtgtgtgt gcgtggggag cgccgcgtgc ggctccgcgc tgcccggcgg ctgtgagcgc
      301 tgcgggcgcg gcgcggggct ttttgcgctc cgcagtgtgc gcgaggggag cgcggccggg
      361 ggcggtgccc cgcggtgcgg ggggggctgc gaggggaaca aaggctgcgt gcggggtgtg
      421 tgcgtggggg ggtgagcagg gggtgtgggc gcgtcggtcg ggctgcaacc ccccctgcac
      481 ccccctcccc gagttgctga gcacggcccg gcttcgggtg cggggctccg tacggggcgt
      541 ggcgcggggc tcgccgtgcc gggcgggggg tggcggcagg tgggggtgcc gggcggggcg
      601 gggccgcctc gggccgggga gggctcgggg gaggggcgcg gcggcccccg gagcgccggc
      661 ggctgtcgag gcgcggcgag ccgcagccat tgccttttat ggtaatcgtg cgagagggcg
      721 cagggacttc ctttgtccca aatctgtgcg gagccgaaat ctgggaggcg ccgccgcacc
      781 ccctctagcg ggcgcggggc gaagcggtgc ggcgccggca ggaaggaaat gggcggggag
      841 ggccttcgtg cgtcgccgcg ccgccgtccc cttctccctc tccagcctcg gggctgtccg
      901 cggggggacg gctgccttcg ggggggacgg ggcagggcgg ggttcggctt ctggcgtgtg
      961 accggcggct ctagagcctc tgctaaccat gttcatgcct tcttcttttt cctacagctc
     1021 ctgggcaacg tgctggttat tgtgctgtct catcattttg gcaaagaatc tagataccct
     1081 cgtaaaggtc tagagctagc ggatcgaacc cttgccacca tggacaagaa gtactccatt
     1141 gggctcgcta tcggcacaaa cagcgtcggc tgggccgtca ttacggacga gtacaaggtg
     1201 ccgagcaaaa aattcaaagt tctgggcaat accgatcgcc acagcataaa gaagaacctc
     1261 attggcgccc tcctgttcga ctccggggag acggccgaag ccacgcggct caaaagaaca
     1321 gcacggcgca gatatacccg cagaaagaat cggatctgct acctgcagga gatctttagt
     1381 aatgagatgg ctaaggtgga tgactctttc ttccataggc tggaggagtc ctttttggtg
     1441 gaggaggata aaaagcacga gcgccaccca atctttggca atatcgtgga cgaggtggcg
     1501 taccatgaaa agtacccaac catatatcat ctgaggaaga agcttgtaga cagtactgat
     1561 aaggctgact tgcggttgat ctatctcgcg ctggcgcata tgatcaaatt tcggggacac
     1621 ttcctcatcg agggggacct gaacccagac aacagcgatg tcgacaaact ctttatccaa
     1681 ctggttcaga cttacaatca gcttttcgaa gagaacccga tcaacgcatc cggagttgac
     1741 gccaaagcaa tcctgagcgc taggctgtcc aaatcccggc ggctcgaaaa cctcatcgca
     1801 cagctccctg gggagaagaa gaacggcctg tttggtaatc ttatcgccct gtcactcggg
     1861 ctgaccccca actttaaatc taacttcgac ctggccgaag atgccaagct tcaactgagc
     1921 aaagacacct acgatgatga tctcgacaat ctgctggccc agatcggcga ccagtacgca
     1981 gacctttttt tggcggcaaa gaacctgtca gacgccattc tgctgagtga tattctgcga
     2041 gtgaacacgg agatcaccaa agctccgctg agcgctagta tgatcaagcg ctatgatgag
     2101 caccaccaag acttgacttt gctgaaggcc cttgtcagac agcaactgcc tgagaagtac
     2161 aaggaaattt tcttcgatca gtctaaaaat ggctacgccg gatacattga cggcggagca
     2221 agccaggagg aattttacaa atttattaag cccatcttgg aaaaaatgga cggcaccgag
     2281 gagctgctgg taaagcttaa cagagaagat ctgttgcgca aacagcgcac tttcgacaat
     2341 ggaagcatcc cccaccagat tcacctgggc gaactgcacg ctatcctcag gcggcaagag
     2401 gatttctacc cctttttgaa agataacagg gaaaagattg agaaaatcct cacatttcgg
     2461 ataccctact atgtaggccc cctcgcccgg ggaaattcca gattcgcgtg gatgactcgc
     2521 aaatcagaag agaccatcac tccctggaac ttcgaggaag tcgtggataa gggggcctct
     2581 gcccagtcct tcatcgaaag gatgactaac tttgataaaa atctgcctaa cgaaaaggtg
     2641 cttcctaaac actctctgct gtacgagtac ttcacagttt ataacgagct caccaaggtc
     2701 aaatacgtca cagaagggat gagaaagcca gcattcctgt ctggagagca gaagaaagct
     2761 atcgtggacc tcctcttcaa gacgaaccgg aaagttaccg tgaaacagct caaagaagac
     2821 tatttcaaaa agattgaatg tttcgactct gttgaaatca gcggagtgga ggatcgcttc
     2881 aacgcatccc tgggaacgta tcacgatctc ctgaaaatca ttaaagacaa ggacttcctg
     2941 gacaatgagg agaacgagga cattcttgag gacattgtcc tcacccttac gttgtttgaa
     3001 gatagggaga tgattgaaga acgcttgaaa acttacgctc atctcttcga cgacaaagtc
     3061 atgaaacagc tcaagaggcg ccgatataca ggatgggggc ggctgtcaag aaaactgatc
     3121 aatgggatcc gagacaagca gagtggaaag acaatcctgg attttcttaa gtccgatgga
     3181 tttgccaacc ggaacttcat gcagttgatc catgatgact ctctcacctt taaggaggac
     3241 atccagaaag cacaagtttc tggccagggg gacagtcttc acgagcacat cgctaatctt
     3301 gcaggtagcc cagctatcaa aaagggaata ctgcagaccg ttaaggtcgt ggatgaactc
     3361 gtcaaagtaa tgggaaggca taagcccgag aatatcgtta tcgagatggc ccgagagaac
     3421 caaactaccc agaagggaca gaagaacagt agggaaagga tgaagaggat tgaagagggt
     3481 ataaaagaac tggggtccca aatccttaag gaacacccag ttgaaaacac ccagcttcag
     3541 aatgagaagc tctacctgta ctacctgcag aacggcaggg acatgtacgt ggatcaggaa
     3601 ctggacatca atcggctctc cgactacgac gtggctgcta tcgtgcccca gtcttttctc
     3661 aaagatgatt ctattgataa taaagtgttg acaagatccg ataaagctag agggaagagt
     3721 gataacgtcc cctcagaaga agttgtcaag aaaatgaaaa attattggcg gcagctgctg
     3781 aacgccaaac tgatcacaca acggaagttc gataatctga ctaaggctga acgaggtggc
     3841 ctgtctgagt tggataaagc cggcttcatc aaaaggcagc ttgttgagac acgccagatc
     3901 accaagcacg tggcccaaat tctcgattca cgcatgaaca ccaagtacga tgaaaatgac
     3961 aaactgattc gagaggtgaa agttattact ctgaagtcta agctggtctc agatttcaga
     4021 aaggactttc agttttataa ggtgagagag atcaacaatt accaccatgc gcatgatgcc
     4081 tacctgaatg cagtggtagg cactgcactt atcaaaaaat atcccaagct tgaatctgaa
     4141 tttgtttacg gagactataa agtgtacgat gttaggaaaa tgatcgcaaa gtctgagcag
     4201 gaaataggca aggccaccgc taagtacttc ttttacagca atattatgaa ttttttcaag
     4261 accgagatta cactggccaa tggagagatt cggaagcgac cacttatcga aacaaacgga
     4321 gaaacaggag aaatcgtgtg ggacaagggt agggatttcg cgacagtccg gaaggtcctg
     4381 tccatgccgc aggtgaacat cgttaaaaag accgaagtac agaccggagg cttctccaag
     4441 gaaagtatcc tcccgaaaag gaacagcgac aagctgatcg cacgcaaaaa agattgggac
     4501 cccaagaaat acggcggatt cgattctcct acagtcgctt acagtgtact ggttgtggcc
     4561 aaagtggaga aagggaagtc taaaaaactc aaaagcgtca aggaactgct gggcatcaca
     4621 atcatggagc gatcaagctt cgaaaaaaac cccatcgact ttctcgaggc gaaaggatat
     4681 aaagaggtca aaaaagacct catcattaag cttcccaagt actctctctt tgagcttgaa
     4741 aacggccgga aacgaatgct cgctagtgcg ggcgagctgc agaaaggtaa cgagctggca
     4801 ctgccctcta aatacgttaa tttcttgtat ctggccagcc actatgaaaa gctcaaaggg
     4861 tctcccgaag ataatgagca gaagcagctg ttcgtggaac aacacaaaca ctaccttgat
     4921 gagatcatcg agcaaataag cgaattctcc aaaagagtga tcctcgccga cgctaacctc
     4981 gataaggtgc tttctgctta caataagcac agggataagc ccatcaggga gcaggcagaa
     5041 aacattatcc acttgtttac tctgaccaac ttgggcgcgc ctgcagcctt caagtacttc
     5101 gacaccacca tagacagaaa gcggtacacc tctacaaagg aggtcctgga cgccacactg
     5161 attcatcagt caattacggg gctctatgaa acaagaatcg acctctctca gctcggtgga
     5221 gacagcaggg ctgaccccaa gaagaagagg aaggtgagtg gtggaggaag tggcgggtca
     5281 gggtcgatgg acgcgaaatc acttacggca tggtcgagaa cactggttac gttcaaggac
     5341 gtgtttgtgg actttacacg tgaggagtgg aaattgctgg atactgcgca acaaattgtg
     5401 tatcgaaatg tcatgcttga gaattacaag aacctcgtca gtctcggata ccagttgacg
     5461 aaaccggatg tgatccttag gctcgaaaag ggggaagaac cttggctggt atcgggaggt
     5521 ggttcgggtg gctctggatc aagcccaaag aagaaacgga aggtggaagc ctcagtgcag
     5581 gtgaaaaggg tgctggaaaa atcccccggc aaactcctcg tgaagatgcc cttccaggct
     5641 tcccctggcg gaaaaggtga agggggtggc gcaaccacat ctgcccaggt catggtcatc
     5701 aagcgacctg gaaggaaaag aaaggccgag gctgaccctc aggccattcc aaagaaacgg
     5761 ggacgcaagc cagggtccgt ggtcgcagct gcagcagctg aggctaagaa aaaggcagtg
     5821 aaggaaagct ccatccgcag tgtgcaggag actgtcctgc ccatcaagaa gaggaagact
     5881 agggagaccg tgtccatcga ggtcaaagaa gtggtcaagc ccctgctcgt gtccaccctg
     5941 ggcgaaaaat ctggaaaggg gctcaaaaca tgcaagtcac ctggacggaa aagcaaggag
     6001 tctagtccaa aggggcgctc aagctccgct tctagtcccc ctaaaaagga acaccatcac
     6061 catcaccatc acgccgagtc tcctaaggct cctatgccac tgctcccacc acctccacca
     6121 cctgagccac agtcaagcga agaccccatc agcccacccg agcctcagga tctgtcctct
     6181 agtatttgca aagaggaaaa gatgcccaga gcaggcagcc tggagagtga tggctgtcca
     6241 aaagaacccg ccaagaccca gcctatggtg gcagccgctg caactaccac cacaaccaca
     6301 actaccacag tggccgaaaa atacaagcat cgcggcgagg gcgaacgaaa ggacattgtg
     6361 tcaagctcca tgcccagacc taaccgggag gaaccagtcg atagtaggac acccgtgact
     6421 gagagagtct catagaagaa ttcgatccct accggttagt aatgagttta aacgggggag
     6481 gctaactgaa acacggaagg agacaatacc ggaaggaacc cgcgctatga cggcaataaa
     6541 aagacagaat aaaacgcacg ggtgttgggt cgtttgttca taaacgcggg gttcggtccc
     6601 agggctggca ctctgtcgat accccaccga gaccccattg gggccaatac gcccgcgttt
     6661 cttccttttc cccaccccac cccccaagtt cgggtgaagg cccagggctc gcagccaacg
     6721 tcggggcggc aggccctgcc atagcagatc tgcgaaatcg gatccgcggc cgcactgatt
     6781 ctagctgcag aggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg
     6841 agaagttggg gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa
     6901 actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt
     6961 atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac
     7021 aggatcctcg agccaccatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg
     7081 agggctccgt gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg
     7141 agggcaccca gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg
     7201 acatcctgtc ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca
     7261 tccccgacta cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact
     7321 tcgaggacgg cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca
     7381 tctacaaggt gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga
     7441 agaccatggg ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg
     7501 gcgagatcaa gcagaggctg aagctgaagg acggcggcca ctacgacgct gaggtcaaga
     7561 ccacctacaa ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagt
     7621 tggacatcac ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg
     7681 gccgccactc caccggcggc atggacgagc tgtacaagta agttaacttg tttattgcag
     7741 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
     7801 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctggaatt
     7861 gactcaaatg atgtcaatta gtctatcaga agctcatctg gtctcccttc cgggggacaa
     7921 gacatccctg tttaatattt aaacagcagt gttcccaaac tgggttctta tatcccttgc
     7981 tctggtcaac caggttgcag ggtttcctgt cctcacagga acgaagtccc taaagaaaca
     8041 gtggcagcca ggtttagccc cggaattgac tggattcctt ttttagggcc cattggtatg
     8101 gctttttccc cgtatccccc caggtgtctg caggctcaaa gagcagcgag aagcgttcag
     8161 aggaaagcga tcccgtgcca ccttccccgt gcccgggctg tccccgcacg ctgccggctc
     8221 ggggatgcgg ggggagcgcc ggaccggagc ggagccccgg gcggctcgct gctgccccct
     8281 agcgggggag ggacgtaatt acatccctgg gggctttggg ggggggctgt ccctgatatc
     8341 tataacaaga aaatatatat ataataagtt atcacgtaag tagaacatga aataacaata
     8401 taattatcgt atgagttaaa tcttaaaagt cacgtaaaag ataatcatgc gtcattttga
     8461 ctcacgcggt cgttatagtt caaaatcagt gacacttacc gcattgacaa gcacgcctca
     8521 cgggagctcc aagcggcgac tgagatgtcc taaatgcaca gcgacggatt cgcgctattt
     8581 agaaagagag agcaatattt caagaatgca tgcgtcaatt ttacgcagac tatctttcta
     8641 gggttaatct agctgcatca ggatcatatc gtcgggtctt ttttccggct cagtcatcgc
     8701 ccaagctggc gctatctggg catcggggag gaagaagccc gtgccttttc ccgcgaggtt
     8761 gaagcggcat ggaaagagtt tgccgaggat gactgctgct gcattgacgt tgagcgaaaa
     8821 cgcacgttta ccatgatgat tcgggaaggt gtggccatgc acgcctttaa cggtgaactg
     8881 ttcgttcagg ccacctggga taccagttcg tcgcggcttt tccggacaca gttccggatg
     8941 gtcagcccga agcgcatcag caacccgaac aataccggcg acagccggaa ctgccgtgcc
     9001 ggtgtgcaga ttaatgacag cggtgcggcg ctgggatatt acgtcagcga ggacgggtat
     9061 cctggctgga tgccgcagaa atggacatgg ataccccgtg agttacccgg cgggcgcgct
     9121 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     9181 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     9241 tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc
     9301 tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg
     9361 cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc
     9421 actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt
     9481 gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc
     9541 ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa
     9601 acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc
     9661 ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg
     9721 cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc
     9781 tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc
     9841 gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca
     9901 ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact
     9961 acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg
    10021 gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt
    10081 ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct
    10141 tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga
    10201 gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa
    10261 tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac
    10321 ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga
    10381 taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc
    10441 cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca
    10501 gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta
    10561 gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg
    10621 tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc
    10681 gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg
    10741 ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt
    10801 ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt
    10861 cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata
    10921 ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc
    10981 gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac
    11041 ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa
    11101 ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct
    11161 tcctttttca atattattga agcatttatc agggttattg tctcatgagc ggatacatat
    11221 ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc
    11281 cacctaaatt gtaagcgtta atattttgtt aaaattcgcg ttaaattttt gttaaatcag
    11341 ctcatttttt aaccaatagg ccgaaatcgg caaaatccct tataaatcaa aagaatagac
    11401 cgagataggg ttgagtgttg ttccagtttg gaacaagagt ccactattaa agaacgtgga
    11461 ctccaacgtc aaagggcgaa aaaccgtcta tcagggcgat ggcccactac gtgaaccatc
    11521 accctaatca agttttttgg ggtcgaggtg ccgtaaagca ctaaatcgga accctaaagg
    11581 gagcccccga tttagagctt gacggggaaa gccggcgaac gtggcgagaa aggaagggaa
    11641 gaaagcgaaa ggagcgggcg ctagggcgct ggcaagtgta gcggtcacgc tgcgcgtaac
    11701 caccacaccc gccgcgctta atgcgccgct acagggcgcg tcccattcgc cattcaggct
    11761 gcgcaactgt tgggaagggc gatcggtgcg ggcctcttcg ctattacgcc agctggcgaa
    11821 agggggatgt gctgcaaggc gattaagttg ggtaacgcca gggttttccc agtcacgacg
    11881 ttgtaaaacg acggccagtg agcgcgcctc gttcattcac gtttttgaac ccgtggagga
    11941 cgggcagact cgcggtgcaa atgtgtttta cagcgtgatg gagcagatga agatgctcga
    12001 cacgctgcag aacacgcagc tagattaacc ctagaaagat aatcatattg tgacgtacgt
    12061 taaagataat catgtgtaaa attgacgcat gtgttttatc ggtctgtata tcgaggttta
    12121 tttattaatt tgaatagata ttaagtttta ttatatttac acttacatac taataataaa
    12181 ttcaacaaac aatttattta tgtttattta tttattaaaa aaaacaaaaa ctcaaaattt
    12241 cttctataaa gtaacaaaac ttttatgagg gacagccccc ccccaaagcc cccagggatg
    12301 taattacgtc cctcccccgc tagggggcag cagcgagccg cccggggctc cgctccggtc
    12361 cggcgctccc cccgcatccc cgagccggca gcgtgcgggg acagcccggg cacggggaag
    12421 gtggcacggg atcgctttcc tctgaacgct tctcgctgct ctttgagcct gcagacacct
    12481 ggggggatac ggggaaaagg cctccacggc cgacattgat tattgactag ttattaatag
    12541 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
    12601 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
    12661 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
    12721 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
    12781 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
    12841 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg
    12901 agccccacgt tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat
    12961 ttatttattt tttaattatt ttgtgcagcg atgggggcgg gggggggggg ggggcgcgcc
    13021 rggggggggg gggggggggg gggggggggg sggggsgrgg cggagaggtg cggcggcagc
    13081 caatcagagc ggcgcgctcc gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc
    13141 ctataaaaag cgaagcgcgc ggcgggcg
//
