LOCUS       sequence-256238-       11360 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  targeted DNA demethylation_mouse_TSDR, expression of 
            dCas9-huTET1CD-T2A-mCherry and sgRNA1 targeting mouse TSDR .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11360)
  AUTHORS   Kressler C, Gasparoni G, Nordstrom K, Hamo D, Salhab A, 
            Dimitropoulos C, Tierling S, Reinke P, Volk HD, Walter J, Hamann A, 
            Polansky JK
  TITLE     Targeted De-Methylation of the FOXP3-TSDR Is Sufficient to Induce 
            Physiological FOXP3 Expression but Not a Functional Treg Phenotype.
  JOURNAL   Front Immunol. 2021 Jan 7;11:609891. doi: 10.3389/fimmu.2020.609891.
            eCollection 2020.
  PUBMED    33488615
REFERENCE   2  (bases 1 to 11360)
  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: "Front 
            Immunol."; date: "2021-01-7"; volume: "11:609891. doi"; pages: " 
            10.3389/fimmu.2020.609891. eCollection 2020"
COMMENT     Sequence Label: pSpdCas9-huTET1CD-T2A-mCherry(PX458)-SgmouseTS...
FEATURES             Location/Qualifiers
     source          1..11360
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        40..288
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     40..60
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     211..230
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        310..385
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        482..767
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     intron          1001..1228
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             1249..1275
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1279..1299
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1309..1332,1333..1335,1336..3891,3892..3894,
                     3895..5409)
                     /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 N863A 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"
     CDS             join(1309..1332,1333..1335,1336..3822,3823..3825,
                     3826..5409)
                     /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"
     CDS             7651..7704
                     /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             7705..8412
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(7847..7865)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(8440..8457)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8446..8653
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   8662..8802
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   8662..8791
                     /label=AAV2 ITR
     rep_origin      8877..9332
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8964..8983)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9174..9195
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9349..9368)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     9468..9490
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(9528..9546)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9614..9718
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9719..9787,9788..10579)
                     /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(9937..9956)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      10750..11338
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11239..11258
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tacggttcct ggccttttgc tggccttttg ctcacatgtg agggcctatt tcccatgatt
       61 ccttcatatt tgcatatacg atacaaggct gttagagaga taattggaat taatttgact
      121 gtaaacacaa agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag
      181 tttgcagttt taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag
      241 tatttcgatt tcttggcttt atatatcttg tggaaaggac gaaacaccgc ttgagattct
      301 aaaatccgtg ttttagagct agaaatagca agttaaaata aggctagtcc gttatcaact
      361 tgaaaaagtg gcaccgagtc ggtgcttttt tgttttagag ctagaaatag caagttaaaa
      421 taaggctagt ccgtttttag cgcgtgcgcc aattctgcag acaaatggct ctagaggtac
      481 ccgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
      541 tgacgtcaat agtaacgcca atagggactt tccattgacg tcaatgggtg gagtatttac
      601 ggtaaactgc ccacttggca gtacatcaag tgtatcatat gccaagtacg ccccctattg
      661 acgtcaatga cggtaaatgg cccgcctggc attgtgccca gtacatgacc ttatgggact
      721 ttcctacttg gcagtacatc tacgtattag tcatcgctat taccatggtc gaggtgagcc
      781 ccacgttctg cttcactctc cccatctccc ccccctcccc acccccaatt ttgtatttat
      841 ttatttttta attattttgt gcagcgatgg gggcgggggg gggggggcga ggcggagagg
      901 tgcggcggca gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg
      961 gcggcggcgg ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgcgctgcct
     1021 tcgccccgtg ccccgctccg ccgccgcctc gcgccgcccg ccccggctct gactgaccgc
     1081 gttactccca caggtgagcg ggcgggacgg cccttctcct ccgggctgta attagctgag
     1141 caagaggtaa gggtttaagg gatggttggt tggtggggta ttaatgttta attacctgga
     1201 gcacctgcct gaaatcactt tttttcaggt tggaccggtg ccaccatgta cccatacgat
     1261 gttccagatt acgcttcgcc gaagaaaaag cgcaaggtcg aagcgtccga caagaagtac
     1321 agcatcggcc tggccatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac
     1381 aaggtgccca gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag
     1441 aacctgatcg gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag
     1501 agaaccgcca gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc
     1561 ttcagcaacg agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc
     1621 ctggtggaag aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag
     1681 gtggcctacc acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc
     1741 accgacaagg ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg
     1801 ggccacttcc tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc
     1861 atccagctgg tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc
     1921 gtggacgcca aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg
     1981 atcgcccagc tgcccggcga gaagaagaat ggcctgttcg gcaacctgat tgccctgagc
     2041 ctgggcctga cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag
     2101 ctgagcaagg acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag
     2161 tacgccgacc tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc
     2221 ctgagagtga acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac
     2281 gacgagcacc accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag
     2341 aagtacaaag agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc
     2401 ggagccagcc aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc
     2461 accgaggaac tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc
     2521 gacaacggca gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg
     2581 caggaagatt tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc
     2641 ttccgcatcc cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg
     2701 accagaaaga gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc
     2761 gcttccgccc agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag
     2821 aaggtgctgc ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc
     2881 aaagtgaaat acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa
     2941 aaggccatcg tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa
     3001 gaggactact tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat
     3061 cggttcaacg cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac
     3121 ttcctggaca atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg
     3181 tttgaggaca gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac
     3241 aaagtgatga agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag
     3301 ctgatcaacg gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc
     3361 gacggcttcg ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa
     3421 gaggacatcc agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc
     3481 aatctggccg gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac
     3541 gagctcgtga aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga
     3601 gagaaccaga ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa
     3661 gagggcatca aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag
     3721 ctgcagaacg agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac
     3781 caggaactgg acatcaaccg gctgtccgac tacgatgtgg acgccatcgt gcctcagagc
     3841 tttctgaagg acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc
     3901 aagagcgaca acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag
     3961 ctgctgaacg ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga
     4021 ggcggcctga gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg
     4081 cagatcacaa agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag
     4141 aatgacaagc tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat
     4201 ttccggaagg atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac
     4261 gacgcctacc tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa
     4321 agcgagttcg tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc
     4381 gagcaggaaa tcggcaaggc taccgccaag tacttcttct acagcaacat catgaacttt
     4441 ttcaagaccg agattaccct ggccaacggc gagatccgga agcggcctct gatcgagaca
     4501 aacggcgaaa ccggggagat cgtgtgggat aagggccggg attttgccac cgtgcggaaa
     4561 gtgctgagca tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc
     4621 agcaaagagt ctatcctgcc caagaggaac agcgataagc tgatcgccag aaagaaggac
     4681 tgggacccta agaagtacgg cggcttcgac agccccaccg tggcctattc tgtgctggtg
     4741 gtggccaaag tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga gctgctgggg
     4801 atcaccatca tggaaagaag cagcttcgag aagaatccca tcgactttct ggaagccaag
     4861 ggctacaaag aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc cctgttcgag
     4921 ctggaaaacg gccggaagag aatgctggcc tctgccggcg aactgcagaa gggaaacgaa
     4981 ctggccctgc cctccaaata tgtgaacttc ctgtacctgg ccagccacta tgagaagctg
     5041 aagggctccc ccgaggataa tgagcagaaa cagctgtttg tggaacagca caagcactac
     5101 ctggacgaga tcatcgagca gatcagcgag ttctccaaga gagtgatcct ggccgacgct
     5161 aatctggaca aagtgctgtc cgcctacaac aagcaccggg ataagcccat cagagagcag
     5221 gccgagaata tcatccacct gtttaccctg accaatctgg gagcccctgc cgccttcaag
     5281 tactttgaca ccaccatcga ccggaagagg tacaccagca ccaaagaggt gctggacgcc
     5341 accctgatcc accagagcat caccggcctg tacgagacac ggatcgacct gtctcagctg
     5401 ggaggcgaca aaaggccggc ggccacgaaa aaggccggcc agggaggagg atccctgccc
     5461 acctgcagct gtcttgatcg agttatacaa aaagacaaag gcccatatta tacacacctt
     5521 ggggcaggac caagtgttgc tgctgtcagg gaaatcatgg agaataggta tggtcaaaaa
     5581 ggaaacgcaa taaggataga aatagtagtg tacaccggta aagaagggaa aagctctcat
     5641 gggtgtccaa ttgctaagtg ggttttaaga agaagcagtg atgaagaaaa agttctttgt
     5701 ttggtccggc agcgtacagg ccaccactgt ccaactgctg tgatggtggt gctcatcatg
     5761 gtgtgggatg gcatccctct tccaatggcc gaccggctat acacagagct cacagagaat
     5821 ctaaagtcat acaatgggca ccctaccgac agaagatgca ccctcaatga aaatcgtacc
     5881 tgtacatgtc aaggaattga tccagagact tgtggagctt cattctcttt tggctgttca
     5941 tggagtatgt actttaatgg ctgtaagttt ggtagaagcc caagccccag aagatttaga
     6001 attgatccaa gctctccctt acatgaaaaa aaccttgaag ataacttaca gagtttggct
     6061 acacgattag ctccaattta taagcagtat gctccagtag cttaccaaaa tcaggtggaa
     6121 tatgaaaatg ttgcccgaga atgtcggctt ggcagcaagg aaggtcgacc cttctctggg
     6181 gtcactgctt gcctggactt ctgtgctcat ccccacaggg acattcacaa catgaataat
     6241 ggaagcactg tggtttgtac cttaactcga gaagataacc gctctttggg tgttattcct
     6301 caagatgagc agctccatgt gctacctctt tataagcttt cagacacaga tgagtttggc
     6361 tccaaggaag gaatggaagc caagatcaaa tctggggcca tcgaggtcct ggcaccccgc
     6421 cgcaaaaaaa gaacgtgttt cactcagcct gttccccgtt ctggaaagaa gagggctgcg
     6481 atgatgacag aggttcttgc acataagata agggcagtgg aaaagaaacc tattccccga
     6541 atcaagcgga agaataactc aacaacaaca aacaacagta agccttcgtc actgccaacc
     6601 ttagggagta acactgagac cgtgcaacct gaagtaaaaa gtgaaaccga accccatttt
     6661 atcttaaaaa gttcagacaa cactaaaact tattcgctga tgccatccgc tcctcaccca
     6721 gtgaaagagg catctccagg cttctcctgg tccccgaaga ctgcttcagc cacaccagct
     6781 ccactgaaga atgacgcaac agcctcatgc gggttttcag aaagaagcag cactccccac
     6841 tgtacgatgc cttcgggaag actcagtggt gccaatgctg cagctgctga tggccctggc
     6901 atttcacagc ttggcgaagt ggctcctctc cccaccctgt ctgctcctgt gatggagccc
     6961 ctcattaatt ctgagccttc cactggtgtg actgagccgc taacgcctca tcagccaaac
     7021 caccagccct ccttcctcac ctctcctcaa gaccttgcct cttctccaat ggaagaagat
     7081 gagcagcatt ctgaagcaga tgagcctcca tcagacgaac ccctatctga tgaccccctg
     7141 tcacctgctg aggagaaatt gccccacatt gatgagtatt ggtcagacag tgagcacatc
     7201 tttttggatg caaatattgg tggggtggcc atcgcacctg ctcacggctc ggttttgatt
     7261 gagtgtgccc ggcgagagct gcacgctacc actcctgttg agcaccccaa ccgtaatcat
     7321 ccaacccgcc tctcccttgt cttttaccag cacaaaaacc taaataagcc ccaacatggt
     7381 tttgaactaa acaagattaa gtttgaggct aaagaagcta agaataagaa aatgaaggcc
     7441 tcagagcaaa aagaccaggc agctaatgaa ggtccagaac agtcctctga agtaaatgaa
     7501 ttgaaccaaa ttccttctca taaagcatta acattaaccc atgacaatgt tgtcaccgtg
     7561 tccccttatg ctctcacaca cgttgcgggg ccctataacc attgggtcat ggccggccag
     7621 gcaaaaaaga aaaaggaatt cggcagtgga gagggcagag gaagtctgct aacatgcggt
     7681 gacgtcgagg agaatcctgg cccagtgagc aagggcgagg aggataacat ggccatcatc
     7741 aaggagttca tgcgcttcaa ggtgcacatg gagggctccg tgaacggcca cgagttcgag
     7801 atcgagggcg agggcgaggg ccgcccctac gagggcaccc agaccgccaa gctgaaggtg
     7861 accaagggtg gccccctgcc cttcgcctgg gacatcctgt cccctcagtt catgtacggc
     7921 tccaaggcct acgtgaagca ccccgccgac atccccgact acttgaagct gtccttcccc
     7981 gagggcttca agtgggagcg cgtgatgaac ttcgaggacg gcggcgtggt gaccgtgacc
     8041 caggactcct ccctgcagga cggcgagttc atctacaagg tgaagctgcg cggcaccaac
     8101 ttcccctccg acggccccgt aatgcagaag aaaaccatgg gctgggaggc ctcctccgag
     8161 cggatgtacc ccgaggacgg cgccctgaag ggcgagatca agcagaggct gaagctgaag
     8221 gacggcggcc actacgacgc tgaggtcaag accacctaca aggccaagaa gcccgtgcag
     8281 ctgcccggcg cctacaacgt caacatcaag ttggacatca cctcccacaa cgaggactac
     8341 accatcgtgg aacagtacga acgcgccgag ggccgccact ccaccggcgg catggacgag
     8401 ctgtacaagt aagaattcta actagagctc gctgatcagc ctcgactgtg ccttctagtt
     8461 gccagccatc tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc
     8521 ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt
     8581 ctattctggg gggtggggtg gggcaggaca gcaaggggga ggattgggaa gagaatagca
     8641 ggcatgctgg ggagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     8701 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     8761 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt
     8821 ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc
     8881 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     8941 acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     9001 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     9061 tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
     9121 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     9181 cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg
     9241 gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     9301 gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc
     9361 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     9421 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     9481 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     9541 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     9601 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     9661 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     9721 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     9781 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     9841 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     9901 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     9961 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
    10021 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
    10081 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
    10141 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
    10201 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
    10261 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
    10321 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
    10381 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg
    10441 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
    10501 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
    10561 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
    10621 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
    10681 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
    10741 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
    10801 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
    10861 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
    10921 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
    10981 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
    11041 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
    11101 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
    11161 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
    11221 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
    11281 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
    11341 cgccagcaac gcggcctttt
//
