LOCUS       sequence-256113-       11365 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  targeted DNA demethylation_mouse_TSDR, expression of 
            dCas9-huTET1CD-T2A-mCherry and sgRNA5 targeting mouse TSDR .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 11365)
  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 11365)
  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..11365
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        68..316
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     68..88
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     239..258
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        338..413
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        510..795
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     intron          1034..1261
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             1282..1308
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1312..1332
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1342..1365,1366..1368,1369..3924,3925..3927,
                     3928..5442)
                     /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(1342..1365,1366..1368,1369..3855,3856..3858,
                     3859..5442)
                     /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             7684..7737
                     /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             7738..8445
                     /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(7880..7898)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(8473..8490)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8479..8686
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   8695..8835
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   8695..8824
                     /label=AAV2 ITR
     rep_origin      8910..9365
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8997..9016)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9207..9228
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9382..9401)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     9501..9523
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(9561..9579)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        9647..9751
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(9752..9820,9821..10612)
                     /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(9970..9989)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      join(10783..11365,1..6)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11272..11291
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
       61 cacatgtgag ggcctatttc ccatgattcc ttcatatttg catatacgat acaaggctgt
      121 tagagagata attggaatta atttgactgt aaacacaaag atattagtac aaaatacgtg
      181 acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga
      241 ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat atatcttgtg
      301 gaaaggacga aacaccgtgg gtgctagcgg atgtggtgtt ttagagctag aaatagcaag
      361 ttaaaataag gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg
      421 ttttagagct agaaatagca agttaaaata aggctagtcc gtttttagcg cgtgcgccaa
      481 ttctgcagac aaatggctct agaggtaccc gttacataac ttacggtaaa tggcccgcct
      541 ggctgaccgc ccaacgaccc ccgcccattg acgtcaatag taacgccaat agggactttc
      601 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
      661 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
      721 tgtgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
      781 atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc
      841 ccctccccac ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg
      901 gcgggggggg cggggcgggg cgaggcggag aggtgcggcg gcagccaatc agagcggcgc
      961 gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata aaaagcgaag
     1021 cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct ccgccgccgc
     1081 ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga gcgggcggga
     1141 cggcccttct cctccgggct gtaattagct gagcaagagg taagggttta agggatggtt
     1201 ggttggtggg gtattaatgt ttaattacct ggagcacctg cctgaaatca ctttttttca
     1261 ggttggaccg gtgccaccat gtacccatac gatgttccag attacgcttc gccgaagaaa
     1321 aagcgcaagg tcgaagcgtc cgacaagaag tacagcatcg gcctggccat cggcaccaac
     1381 tctgtgggct gggccgtgat caccgacgag tacaaggtgc ccagcaagaa attcaaggtg
     1441 ctgggcaaca ccgaccggca cagcatcaag aagaacctga tcggagccct gctgttcgac
     1501 agcggcgaaa cagccgaggc cacccggctg aagagaaccg ccagaagaag atacaccaga
     1561 cggaagaacc ggatctgcta tctgcaagag atcttcagca acgagatggc caaggtggac
     1621 gacagcttct tccacagact ggaagagtcc ttcctggtgg aagaggataa gaagcacgag
     1681 cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa gtaccccacc
     1741 atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct gcggctgatc
     1801 tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga gggcgacctg
     1861 aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac ctacaaccag
     1921 ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat cctgtctgcc
     1981 agactgagca agagcagacg gctggaaaat ctgatcgccc agctgcccgg cgagaagaag
     2041 aatggcctgt tcggcaacct gattgccctg agcctgggcc tgacccccaa cttcaagagc
     2101 aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta cgacgacgac
     2161 ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct ggccgccaag
     2221 aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga gatcaccaag
     2281 gcccccctga gcgcctctat gatcaagaga tacgacgagc accaccagga cctgaccctg
     2341 ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt cttcgaccag
     2401 agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga gttctacaag
     2461 ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt gaagctgaac
     2521 agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc ccaccagatc
     2581 cacctgggag agctgcacgc cattctgcgg cggcaggaag atttttaccc attcctgaag
     2641 gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta cgtgggccct
     2701 ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga aaccatcacc
     2761 ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt catcgagcgg
     2821 atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca cagcctgctg
     2881 tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac cgagggaatg
     2941 agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct gctgttcaag
     3001 accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa aatcgagtgc
     3061 ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct gggcacatac
     3121 cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga aaacgaggac
     3181 attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat gatcgaggaa
     3241 cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct gaagcggcgg
     3301 agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg ggacaagcag
     3361 tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag aaacttcatg
     3421 cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc
     3481 ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc cgccattaag
     3541 aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat gggccggcac
     3601 aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca gaagggacag
     3661 aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct gggcagccag
     3721 atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct gtacctgtac
     3781 tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc
     3841 gactacgatg tggacgccat cgtgcctcag agctttctga aggacgactc catcgacaac
     3901 aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag
     3961 gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct gattacccag
     4021 agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact ggataaggcc
     4081 ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt ggcacagatc
     4141 ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg ggaagtgaaa
     4201 gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca gttttacaaa
     4261 gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga
     4321 accgccctga tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag
     4381 gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc
     4441 aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac cctggccaac
     4501 ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg
     4561 gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca agtgaatatc
     4621 gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct gcccaagagg
     4681 aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta cggcggcttc
     4741 gacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc
     4801 aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag aagcagcttc
     4861 gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa aaaggacctg
     4921 atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa gagaatgctg
     4981 gcctctgccg gcgaactgca gaagggaaac gaactggccc tgccctccaa atatgtgaac
     5041 ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga taatgagcag
     5101 aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga gcagatcagc
     5161 gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct gtccgcctac
     5221 aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca cctgtttacc
     5281 ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat cgaccggaag
     5341 aggtacacca gcaccaaaga ggtgctggac gccaccctga tccaccagag catcaccggc
     5401 ctgtacgaga cacggatcga cctgtctcag ctgggaggcg acaaaaggcc ggcggccacg
     5461 aaaaaggccg gccagggagg aggatccctg cccacctgca gctgtcttga tcgagttata
     5521 caaaaagaca aaggcccata ttatacacac cttggggcag gaccaagtgt tgctgctgtc
     5581 agggaaatca tggagaatag gtatggtcaa aaaggaaacg caataaggat agaaatagta
     5641 gtgtacaccg gtaaagaagg gaaaagctct catgggtgtc caattgctaa gtgggtttta
     5701 agaagaagca gtgatgaaga aaaagttctt tgtttggtcc ggcagcgtac aggccaccac
     5761 tgtccaactg ctgtgatggt ggtgctcatc atggtgtggg atggcatccc tcttccaatg
     5821 gccgaccggc tatacacaga gctcacagag aatctaaagt catacaatgg gcaccctacc
     5881 gacagaagat gcaccctcaa tgaaaatcgt acctgtacat gtcaaggaat tgatccagag
     5941 acttgtggag cttcattctc ttttggctgt tcatggagta tgtactttaa tggctgtaag
     6001 tttggtagaa gcccaagccc cagaagattt agaattgatc caagctctcc cttacatgaa
     6061 aaaaaccttg aagataactt acagagtttg gctacacgat tagctccaat ttataagcag
     6121 tatgctccag tagcttacca aaatcaggtg gaatatgaaa atgttgcccg agaatgtcgg
     6181 cttggcagca aggaaggtcg acccttctct ggggtcactg cttgcctgga cttctgtgct
     6241 catccccaca gggacattca caacatgaat aatggaagca ctgtggtttg taccttaact
     6301 cgagaagata accgctcttt gggtgttatt cctcaagatg agcagctcca tgtgctacct
     6361 ctttataagc tttcagacac agatgagttt ggctccaagg aaggaatgga agccaagatc
     6421 aaatctgggg ccatcgaggt cctggcaccc cgccgcaaaa aaagaacgtg tttcactcag
     6481 cctgttcccc gttctggaaa gaagagggct gcgatgatga cagaggttct tgcacataag
     6541 ataagggcag tggaaaagaa acctattccc cgaatcaagc ggaagaataa ctcaacaaca
     6601 acaaacaaca gtaagccttc gtcactgcca accttaggga gtaacactga gaccgtgcaa
     6661 cctgaagtaa aaagtgaaac cgaaccccat tttatcttaa aaagttcaga caacactaaa
     6721 acttattcgc tgatgccatc cgctcctcac ccagtgaaag aggcatctcc aggcttctcc
     6781 tggtccccga agactgcttc agccacacca gctccactga agaatgacgc aacagcctca
     6841 tgcgggtttt cagaaagaag cagcactccc cactgtacga tgccttcggg aagactcagt
     6901 ggtgccaatg ctgcagctgc tgatggccct ggcatttcac agcttggcga agtggctcct
     6961 ctccccaccc tgtctgctcc tgtgatggag cccctcatta attctgagcc ttccactggt
     7021 gtgactgagc cgctaacgcc tcatcagcca aaccaccagc cctccttcct cacctctcct
     7081 caagaccttg cctcttctcc aatggaagaa gatgagcagc attctgaagc agatgagcct
     7141 ccatcagacg aacccctatc tgatgacccc ctgtcacctg ctgaggagaa attgccccac
     7201 attgatgagt attggtcaga cagtgagcac atctttttgg atgcaaatat tggtggggtg
     7261 gccatcgcac ctgctcacgg ctcggttttg attgagtgtg cccggcgaga gctgcacgct
     7321 accactcctg ttgagcaccc caaccgtaat catccaaccc gcctctccct tgtcttttac
     7381 cagcacaaaa acctaaataa gccccaacat ggttttgaac taaacaagat taagtttgag
     7441 gctaaagaag ctaagaataa gaaaatgaag gcctcagagc aaaaagacca ggcagctaat
     7501 gaaggtccag aacagtcctc tgaagtaaat gaattgaacc aaattccttc tcataaagca
     7561 ttaacattaa cccatgacaa tgttgtcacc gtgtcccctt atgctctcac acacgttgcg
     7621 gggccctata accattgggt catggccggc caggcaaaaa agaaaaagga attcggcagt
     7681 ggagagggca gaggaagtct gctaacatgc ggtgacgtcg aggagaatcc tggcccagtg
     7741 agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt caaggtgcac
     7801 atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc
     7861 tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct gcccttcgcc
     7921 tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa gcaccccgcc
     7981 gacatccccg actacttgaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg
     8041 aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca ggacggcgag
     8101 ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc cgtaatgcag
     8161 aagaaaacca tgggctggga ggcctcctcc gagcggatgt accccgagga cggcgccctg
     8221 aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga cgctgaggtc
     8281 aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa cgtcaacatc
     8341 aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta cgaacgcgcc
     8401 gagggccgcc actccaccgg cggcatggac gagctgtaca agtaagaatt ctaactagag
     8461 ctcgctgatc agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc
     8521 ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg
     8581 aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg
     8641 acagcaaggg ggaggattgg gaagagaata gcaggcatgc tggggagcgg ccgcaggaac
     8701 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     8761 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     8821 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     8881 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     8941 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
     9001 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     9061 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     9121 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     9181 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     9241 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     9301 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     9361 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     9421 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     9481 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     9541 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     9601 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     9661 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     9721 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     9781 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     9841 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     9901 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     9961 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
    10021 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
    10081 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
    10141 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
    10201 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
    10261 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
    10321 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
    10381 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
    10441 tgataaatct ggagccggtg agcgtggaag ccgcggtatc attgcagcac tggggccaga
    10501 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
    10561 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
    10621 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
    10681 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
    10741 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
    10801 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
    10861 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
    10921 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
    10981 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
    11041 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
    11101 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
    11161 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
    11221 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
    11281 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
    11341 atgctcgtca ggggggcgga gccta
//
