LOCUS       P3-dCas9-Tet1-GF       12956 bp DNA     circular SYN 15-AUG-2022
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    P3-dCas9-Tet1-GFP-Puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12956)
  AUTHORS   sdff
  TITLE     Direct Submission
  JOURNAL   Exported Aug 15, 2022 from SnapGene 6.1.0
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..12956
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..241
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA"
     primer_bind     5..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     267..287
                     /label=Primer 1
     misc_RNA        268..343
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     primer_bind     349..369
                     /label=Primer 1
     enhancer        440..725
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     primer_bind     440..456
                     /label=F EGFP NOTI
     promoter        727..1004
                     /label=chicken beta-actin promoter
     intron          1005..1233
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1245..1254
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1254..1319
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1326..1346
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1371..5471
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     primer_bind     1685..1707
                     /label=F dCAS9
     primer_bind     complement(1817..1836)
                     /label=R dCAS9
     primer_bind     complement(1982..1997)
                     /label=F 3a D
     primer_bind     5428..5446
                     /label=F dcas9-DNMT3a
     CDS             5472..5519
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     5472..5489
                     /label=F 3a D
     CDS             5547..7700
                     /codon_start=1
                     /label=Active Tet1
                     /translation="LPTCSCLDRVIQKDKGPYYTHLGAGPSVAAVREIMENRYGQKGNA
                     IRIEIVVYTGKEGKSSHGCPIAKWVLRRSSDEEKVLCLVRQRTGHHCPTAVMVVLIMVW
                     DGIPLPMADRLYTELTENLKSYNGHPTDRRCTLNENRTCTCQGIDPETCGASFSFGCSW
                     SMYFNGCKFGRSPSPRRFRIDPSSPLHEKNLEDNLQSLATRLAPIYKQYAPVAYQNQVE
                     YENVARECRLGSKEGRPFSGVTACLDFCAHPHRDIHNMNNGSTVVCTLTREDNRSLGVI
                     PQDEQLHVLPLYKLSDTDEFGSKEGMEAKIKSGAIEVLAPRRKKRTCFTQPVPRSGKKR
                     AAMMTEVLAHKIRAVEKKPIPRIKRKNNSTTTNNSKPSSLPTLGSNTETVQPEVKSETE
                     PHFILKSSDNTKTYSLMPSAPHPVKEASPGFSWSPKTASATPAPLKNDATASCGFSERS
                     STPHCTMPSGRLSGANAAAADGPGISQLGEVAPLPTLSAPVMEPLINSEPSTGVTEPLT
                     PHQPNHQPSFLTSPQDLASSPMEEDEQHSEADEPPSDEPLSDDPLSPAEEKLPHIDEYW
                     SDSEHIFLDANIGGVAIAPAHGSVLIECARRELHATTPVEHPNRNHPTRLSLVFYQHKN
                     LNKPQHGFELNKIKFEAKEAKNKKMKASEQKDQAANEGPEQSSEVNELNQIPSHKALTL
                     THDNVVTVSPYALTHVAGPYNHWV"
     primer_bind     complement(5604..5620)
                     /label=qPCR TET1 R
     primer_bind     complement(7691..7709)
                     /label=TET1 reverse cloning
     primer_bind     complement(7736..7753)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    7742..7949
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     7949..7980
                     /label=F EGFP NOTI
     enhancer        7964..8267
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8268..8471
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     regulatory      8510..8519
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             8516..9232
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     9105..9126
                     /label=F sequencing Puro
     primer_bind     9200..9219
                     /label=EGFP sequencing New 2
     primer_bind     9255..9285
                     /label=F puro New stop
     CDS             9263..9316
                     /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             9317..9916
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(9900..9922)
                     /label=R puro New stop
     polyA_signal    10084..10205
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10187..10219)
                     /label=R EGFP NOTI
     repeat_region   10219..10359
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   10219..10348
                     /label=AAV2 ITR
     rep_origin      10434..10889
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10521..10540)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10731..10752
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10906..10925)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     11025..11047
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(11085..11103)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        11171..11275
                     /gene="bla"
                     /label=AmpR promoter
     CDS             11276..12136
                     /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(11494..11513)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      12307..12895
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     12796..12815
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggccgcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg ggtcttcgag aagacctgtt ttagagctag aaatagcaag ttaaaataag
      301 gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg ttttagagct
      361 agaaatagca agttaaaata aggctagtcc gtttttagcg cgtgcgccaa ttctgcagac
      421 aaatggctct agaggtaccc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      481 ccaacgaccc ccgcccattg acgtcaatag taacgccaat agggactttc cattgacgtc
      541 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
      601 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tgtgcccagt
      661 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
      721 ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac
      781 ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg gcgggggggg
      841 ggggggggcg cgcgccaggc ggggcggggc ggggcgaggg gcggggcggg gcgaggcgga
      901 gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa gtttcctttt atggcgaggc
      961 ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg ggcgggagtc gctgcgacgc
     1021 tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg gctctgactg
     1081 accgcgttac tcccacaggt gagcgggcgg gacggccctt ctcctccggg ctgtaattag
     1141 ctgagcaaga ggtaagggtt taagggatgg ttggttggtg gggtattaat gtttaattac
     1201 ctggagcacc tgcctgaaat cacttttttt caggttggac cggtgccacc atggactata
     1261 aggaccacga cggagactac aaggatcatg atattgatta caaagacgat gacgataaga
     1321 tggccccaaa gaagaagcgg aaggtcggta tccacggagt cccagcagcc gacaagaagt
     1381 acagcatcgg cctggccatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt
     1441 acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga
     1501 agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc acccggctga
     1561 agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga
     1621 tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct
     1681 tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg
     1741 aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca
     1801 gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc
     1861 ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt
     1921 tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg
     1981 gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc
     2041 tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggcaacctg attgccctga
     2101 gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc
     2161 agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc
     2221 agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca
     2281 tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat
     2341 acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg
     2401 agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacattgacg
     2461 gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg
     2521 gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct
     2581 tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc
     2641 ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga
     2701 ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga
     2761 tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg
     2821 gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg
     2881 agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat aacgagctga
     2941 ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga
     3001 aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga
     3061 aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag
     3121 atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg
     3181 acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac
     3241 tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg
     3301 acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga
     3361 agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt
     3421 ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta
     3481 aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg
     3541 ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg
     3601 acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca
     3661 gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg
     3721 aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc
     3781 agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg
     3841 accaggaact ggacatcaac cggctgtccg actacgatgt ggacgctatc gtgcctcaga
     3901 gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac aagaaccggg
     3961 gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcggc
     4021 agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc aaggccgaga
     4081 gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc
     4141 ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg
     4201 agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg
     4261 atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc
     4321 acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg
     4381 aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga
     4441 gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact
     4501 ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga
     4561 caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc accgtgcgga
     4621 aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct
     4681 tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc agaaagaagg
     4741 actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg
     4801 tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg
     4861 ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca
     4921 agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg
     4981 agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg
     5041 aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc
     5101 tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaagcact
     5161 acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg
     5221 ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc atcagagagc
     5281 aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca
     5341 agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg
     5401 ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc
     5461 tgggaggcga caaaaggccg gcggccacga aaaaggccgg acaggccaaa aagaaaaagc
     5521 tcgagggcgg aggcgggagc ggatccctgc ccacctgcag ctgtcttgat cgagttatac
     5581 aaaaagacaa aggcccatat tatacacacc ttggggcagg accaagtgtt gctgctgtca
     5641 gggaaatcat ggagaatagg tatggtcaaa aaggaaacgc aataaggata gaaatagtag
     5701 tgtacaccgg taaagaaggg aaaagctctc atgggtgtcc aattgctaag tgggttttaa
     5761 gaagaagcag tgatgaagaa aaagttcttt gtttggtccg gcagcgtaca ggccaccact
     5821 gtccaactgc tgtgatggtg gtgctcatca tggtgtggga tggcatccct cttccaatgg
     5881 ccgaccggct atacacagag ctcacagaga atctaaagtc atacaatggg caccctaccg
     5941 acagaagatg caccctcaat gaaaatcgta cctgtacatg tcaaggaatt gatccagaga
     6001 cttgtggagc ttcattctct tttggctgtt catggagtat gtactttaat ggctgtaagt
     6061 ttggtagaag cccaagcccc agaagattta gaattgatcc aagctctccc ttacatgaaa
     6121 aaaaccttga agataactta cagagtttgg ctacacgatt agctccaatt tataagcagt
     6181 atgctccagt agcttaccaa aatcaggtgg aatatgaaaa tgttgcccga gaatgtcggc
     6241 ttggcagcaa ggaaggtcga cccttctctg gggtcactgc ttgcctggac ttctgtgctc
     6301 atccccacag ggacattcac aacatgaata atggaagcac tgtggtttgt accttaactc
     6361 gagaagataa ccgctctttg ggtgttattc ctcaagatga gcagctccat gtgctacctc
     6421 tttataagct ttcagacaca gatgagtttg gctccaagga aggaatggaa gccaagatca
     6481 aatctggggc catcgaggtc ctggcacccc gccgcaaaaa aagaacgtgt ttcactcagc
     6541 ctgttccccg ttctggaaag aagagggctg cgatgatgac agaggttctt gcacataaga
     6601 taagggcagt ggaaaagaaa cctattcccc gaatcaagcg gaagaataac tcaacaacaa
     6661 caaacaacag taagccttcg tcactgccaa ccttagggag taacactgag accgtgcaac
     6721 ctgaagtaaa aagtgaaacc gaaccccatt ttatcttaaa aagttcagac aacactaaaa
     6781 cttattcgct gatgccatcc gctcctcacc cagtgaaaga ggcatctcca ggcttctcct
     6841 ggtcccccaa gactgcttca gccacaccag ctccactgaa gaatgacgca acagcctcat
     6901 gcgggttttc agaaagaagc agcactcccc actgtacgat gccttcgggt agactcagtg
     6961 gtgccaatgc tgcagctgct gatggccctg gcatttcaca gcttggcgaa gtggctcctc
     7021 tccccaccct gtctgctcct gtgatggagc ccctcattaa ttctgagcct tccactggtg
     7081 tgactgagcc gctaacgcct catcagccaa accaccagcc ctccttcctc acctctcctc
     7141 aagaccttgc ctcttctcca atggaagaag atgagcagca ttctgaagca gatgagcctc
     7201 catcagacga acccctatct gatgaccccc tgtcacctgc tgaggagaaa ttgccccaca
     7261 ttgatgagta ttggtcagac agtgagcaca tctttttgga tgcaaatatt ggtggggtgg
     7321 ccatcgcacc tgctcacggc tcggttttga ttgagtgtgc ccggcgagag ctgcacgcta
     7381 ccactcctgt tgagcacccc aaccgtaatc atccaacccg cctctccctt gtcttttacc
     7441 agcacaaaaa cctaaataag ccccaacatg gttttgaact aaacaagatt aagtttgagg
     7501 ctaaagaagc taagaataag aaaatgaagg cctcagagca aaaagaccag gcagctaatg
     7561 aaggtccaga acagtcctct gaagtaaatg aattgaacca aattccttct cataaagcat
     7621 taacattaac ccatgacaat gttgtcaccg tgtcccctta tgctctcaca cacgttgcgg
     7681 ggccctataa ccattgggtc tgagaattca attctaacta gagctcgctg atcagcctcg
     7741 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     7801 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     7861 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     7921 tgggaagaga atagcaggca tgctggggag cggccgcgtt ccgcgttaca taacttacgg
     7981 taaatggccc gcctggctga ccgcccaacg acccccgccc attgacgtca ataatgacgt
     8041 atgttcccat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
     8101 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
     8161 acgtcaatga cggtaaatgg cccgcctggc attatgccca gtacatgacc ttatgggact
     8221 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtg atgcggtttt
     8281 ggcagtacat caatgggcgt ggatagcggt ttgactcacg gggatttcca agtctccacc
     8341 ccattgacgt caatgggagt ttgttttggc accaaaatca acgggacttt ccaaaatgtc
     8401 gtaacaactc cgccccattg acgcaaatgg gcggtaggcg tgtacggtgg gaggtctata
     8461 taagcagagc tggtttagtg aaccgtcaga tccgctagcg ctaccggtcg ccaccatggt
     8521 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     8581 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     8641 gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     8701 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     8761 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     8821 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     8881 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     8941 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     9001 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     9061 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     9121 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     9181 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtccggact
     9241 cagatctcga gctcaagctt ctgagggcag aggaagtctg ctaacatgcg gtgacgtcga
     9301 ggagaatcct ggcccaatga ccgagtacaa gcccacggtg cgcctcgcca cccgcgacga
     9361 cgtccccagg gccgtacgca ccctcgccgc cgcgttcgcc gactaccccg ccacgcgcca
     9421 caccgtcgat ccggaccgcc acatcgagcg ggtcaccgag ctgcaagaac tcttcctcac
     9481 gcgcgtcggg ctcgacatcg gcaaggtgtg ggtcgcggac gacggcgccg cggtggcggt
     9541 ctggaccacg ccggagagcg tcgaagcggg ggcggtgttc gccgagatcg gcccgcgcat
     9601 ggccgagttg agcggttccc ggctggccgc gcagcaacag atggaaggcc tcctggcgcc
     9661 gcaccggccc aaggagcccg cgtggttcct ggccaccgtc ggcgtctcgc ccgaccacca
     9721 gggcaagggt ctgggcagcg ccgtcgtgct ccccggagtg gaggcggccg agcgcgccgg
     9781 ggtgcccgcc ttcctggaga cctccgcgcc ccacaacctc cccttctacg agcggctcgg
     9841 cttcaccgtc accgccgacg tcgaggtgcc cgaaggaccg cgcacctggt gcatgacccg
     9901 caagcccggt gcctgaaagc ttcgaattct gcagtcgacg gtaccgcggg cccgggatcc
     9961 accggatcta gataactgat cataatcagc cataccacat ttgtagaggt tttacttgct
    10021 ttaaaaaacc tcccacacct ccccctgaac ctgaaacata aaatgaatgc aattgttgtt
    10081 gttaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
    10141 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
    10201 tcttaacgcg gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
    10261 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
    10321 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc
    10381 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
    10441 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
    10501 tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
    10561 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
    10621 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
    10681 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
    10741 gttccaaact ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat
    10801 tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
    10861 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
    10921 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
    10981 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
    11041 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
    11101 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
    11161 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
    11221 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
    11281 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
    11341 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
    11401 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
    11461 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
    11521 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
    11581 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
    11641 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
    11701 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
    11761 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
    11821 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
    11881 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
    11941 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg
    12001 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
    12061 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
    12121 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
    12181 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
    12241 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
    12301 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
    12361 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
    12421 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
    12481 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
    12541 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
    12601 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
    12661 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
    12721 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
    12781 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
    12841 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
    12901 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgt
//
