LOCUS       Exported               10921 bp ds-DNA     circular SYN 27-FEB-2025
DEFINITION  Dox-inducible expression of the engineered epigenetic effector Ezh2 
            fused with scFV-sfGFP to programme H3K27 methylation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10921)
  AUTHORS   Policarpi C, Munafo M, Tsagkris S, Carlini V, Hackett JA.
  TITLE     Systematic epigenome editing captures the context-dependent 
            instructive function of chromatin modifications.
  JOURNAL   Nat Genet 2024 Jun;56:1168-1180
  PUBMED    38724747
REFERENCE   2  (bases 1 to 10921)
  AUTHORS   Epitome Tx
  TITLE     Direct Submission
  JOURNAL   Exported Friday, Feb 28, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
COMMENT     Under patent. Not for commercial usage or modification.
FEATURES             Location/Qualifiers
     source          1..10921
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(131..586)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     728..744
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    complement(870..1104)
                     /label=Piggy 3'
     promoter        1386..1764
                     /label=TRE3G promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G"
     protein_bind    1394..1412
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1430..1448
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1466..1484
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1502..1520
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1538..1556
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1574..1592
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1610..1628
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1711..1730
                     /label=Primer 1
     primer_bind     1763..1798
                     /label=p163_Frag1_F
     CDS             1780..2481
                     /codon_start=1
                     /label=Recoded scFV
                     /translation="MGPDIVMTQSPSSLSASVGDRVTITCRSSTGAVTTSNYASWVQEK
                     PGKLFKGLIGGTNNRAPGVPSRFSGSLIGDKATLTISSLQPEDFATYFCALWYSNHWVF
                     GQGTKVELKRGGGGSGGGGSGGGGSSGGGSEVKLLESGGGLVQPGGSLKLSCAVSGFSL
                     TDYGVNWVRQAPGRGLEWIGVIWGDGITDYNSALKDRFIISKDNGKNTVYLQMSKVRSD
                     DTALYYCVTGLF"
     primer_bind     complement(2171..2190)
                     /label=scFV-recodedgenot_R
     primer_bind     2341..2360
                     /label=Primer 2
     CDS             2521..2547
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2548..2637
                     /codon_start=1
                     /label=Linker
                     /translation="GGGGGSGGGGSGGGGSGGGGSLDPGGGGSG"
     CDS             2638..3348
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     primer_bind     2971..2990
                     /label=Primer 3
     primer_bind     3193..3213
                     /label=OJH_511
     CDS             3367..3387
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3394..3414
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T 
                     antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     misc_feature    3427..3462
                     /label=Linker
     primer_bind     complement(3445..3469)
                     /label=p163_Frag1_R
     primer_bind     3449..3486
                     /label=EZH2_COMPLETECLONINGFORWARD
     exon            3468..3585
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     CDS             3469..5709
                     /codon_start=1
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /product="histone-lysine N-methyltransferase EZH2 isoform
                     1"
                     /EC_number="2.1.1.43"
                     /label=Ezh2 FL
                     /note="isoform 1 is encoded by transcript variant 1; 
                     histone-lysine N-methyltransferase EZH2; enhancer of zeste 
                     homolog 2"
                     /db_xref="CCDS:CCDS20096.1"
                     /db_xref="GeneID:14056"
                     /db_xref="MGI:MGI:107940"
                     /protein_id="NP_031997.2"
                     /translation="MGQTGKKSEKGPVCWRKRVKSEYMRLRQLKRFRRADEVKTMFSSN
                     RQKILERTETLNQEWKQRRIQPVHIMTSVSSLRGTRECSVTSDLDFPAQVIPLKTLNAV
                     ASVPIMYSWSPLQQNFMVEDETVLHNIPYMGDEVLDQDGTFIEELIKNYDGKVHGDREC
                     GFINDEIFVELVNALGQYNDDDDDDDGDDPDEREEKQKDLEDNRDDKETCPPRKFPADK
                     IFEAISSMFPDKGTAEELKEKYKELTEQQLPGALPPECTPNIDGPNAKSVQREQSLHSF
                     HTLFCRRCFKYDCFLHPFHATPNTYKRKNTETALDNKPCGPQCYQHLEGAKEFAAALTA
                     ERIKTPPKRPGGRRRGRLPNNSSRPSTPTISVLESKDTDSDREAGTETGGENNDKEEEE
                     KKDETSSSSEANSRCQTPIKMKPNIEPPENVEWSGAEASMFRVLIGTYYDNFCAIARLI
                     GTKTCRQVYEFRVKESSIIAPVPTEDVDTPPRKKKRKHRLWAAHCRKIQLKKDGSSNHV
                     YNYQPCDHPRQPCDSSCPCVIAQNFCEKFCQCSSECQNRFPGCRCKAQCNTKQCPCYLA
                     VRECDPDLCLTCGAADHWDSKNVSCKNCSIQRGSKKHLLLAPSDVAGWGIFIKDPVQKN
                     EFISEYCGEIISQDEADRRGKVYDKYMCSFLFNLNNDFVVDATRKGNKIRFANHSVNPN
                     CYAKVMMVNGDHRIGIFAKRAIQTGEELFFDYRYSQADALKYVGIEREMEIP"
     misc_feature    3529..3531
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /note="Phosphoserine, by PKB/AKT1. 
                     {ECO:0000250|UniProtKB:Q15910}; propagated from 
                     UniProtKB/Swiss-Prot (Q61188.2); phosphorylation site"
                     /experiment="experimental evidence, no additional details
                     recorded"
     misc_feature    3583..3672
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /note="propagated from UniProtKB/Swiss-Prot (Q61188.2);
                     Region: Interaction with EED"
                     /experiment="experimental evidence, no additional details
                     recorded"
     misc_feature    3583..3669
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /note="WD repeat binding protein EZH2; Region:
                     EZH2_WD-Binding; pfam11616"
                     /db_xref="CDD:288468"
     exon            3586..3714
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     misc_feature    3694..3696
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /note="Phosphoserine. {ECO:0000250|UniProtKB:Q15910};
                     propagated from UniProtKB/Swiss-Prot (Q61188.2); 
                     phosphorylation site"
                     /experiment="experimental evidence, no additional details
                     recorded"
     exon            3715..3831
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     primer_bind     3820..3840
                     /label=p163_seq_Ezh2full_F1
     exon            3832..3952
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            3953..4093
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            4094..4196
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            4197..4360
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            4361..4452
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     primer_bind     4441..4461
                     /label=p163_Ezh2full_seq_F2
     exon            4453..4693
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            4694..4863
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            4864..4958
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     misc_feature    4927..4929
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /note="Phosphothreonine. {ECO:0000244|PubMed:21183079}; 
                     propagated from UniProtKB/Swiss-Prot (Q61188.2); 
                     phosphorylation site"
                     /experiment="experimental evidence, no additional details
                     recorded"
     exon            4959..4999
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            5000..5125
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     primer_bind     5063..5083
                     /label=p163_seq_Ezh2full_F3
     exon            5126..5304
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            5305..5400
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            5401..5482
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            5483..5563
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     exon            5564..5648
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     misc_feature    5644..5646
                     /label=Y to D for catalytic mutant
     exon            5649..5711
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /inference="alignment:same species:1.39.8"
     STS             5657..5711
                     /gene="Ezh2"
                     /gene_synonym="Enx-1; Enx1h; KMT6; mKIAA4065"
                     /standard_name="AW554728"
                     /label=AW554728
                     /db_xref="UniSTS:207946"
     primer_bind     complement(5685..5724)
                     /label=EZH2_COMPLETECLONINGREVERSE
     primer_bind     5710..5743
                     /label=p163_Frag2_F
     misc_feature    5713..7488
     primer_bind     5753..5772
                     /label=Primer 4
     primer_bind     complement(5818..5835)
                     /label=p163_Frag2_R
     polyA_signal    5821..5942
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    5953..7488
     misc_feature    5953..7488
     misc_feature    5957..6477
                     /label=human beta actin promoter
     misc_feature    6232..6251
                     /label=bact-fwd
     misc_feature    complement(6232..6251)
                     /label=bact-rev
     misc_feature    6353..6372
                     /label=betact5
     primer_bind     6383..6402
                     /label=Primer 5
     CDS             6503..7297
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     misc_feature    6628..6647
                     /label=neoseq-fwd
     misc_feature    6672..6691
                     /label=neoPstI_5
     misc_feature    complement(6738..6757)
                     /label=neoseq-rev
     misc_feature    complement(6738..6757)
                     /label=neoseq-rev
     misc_feature    7009..7027
                     /label=neoSphI_5
     primer_bind     7013..7032
                     /label=Primer 6
     misc_feature    7167..7186
                     /label=neoseq2-fwd
     misc_feature    7302..7321
                     /label=neoBstB1_5
     primer_bind     complement(7466..7506)
                     /label=p163_Frag2_R
     polyA_signal    7492..7613
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    7672..7692
                     /label=514
     misc_feature    complement(8104..8413)
                     /label='Piggy 5'
     primer_bind     complement(8900..8916)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8924..8940
                     /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(8948..8978)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8993..9014
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(9302..9890)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(10061..10921)
                     /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"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa
      181 atcagctcat tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa
      241 tagaccgaga tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac
      301 gtggactcca acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa
      361 ccatcaccct aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct
      421 aaagggagcc cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa
      481 gggaagaaag cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc
      541 gtaaccacca cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc
      601 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      661 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      721 cgacgttgta aaacgacggc cagtgagcgc gcctcgttca ttcacgtttt tgaacccgtg
      781 gaggacgggc agactcgcgg tgcaaatgtg ttttacagcg tgatggagca gatgaagatg
      841 ctcgacacgc tgcagaacac gcagctagat taaccctaga aagataatca tattgtgacg
      901 tacgttaaag ataatcatgc gtaaaattga cgcatgtgtt ttatcggtct gtatatcgag
      961 gtttatttat taatttgaat agatattaag ttttattata tttacactta catactaata
     1021 ataaattcaa caaacaattt atttatgttt atttatttat taaaaaaaaa caaaaactca
     1081 aaatttcttc tataaagtaa caaaactttt atgagggaca gccccccccc aaagccccca
     1141 gggatgtaat tacgtccctc ccccgctagg gggcagcagc gagccgcccg gggctccgct
     1201 ccggtccggc gctccccccg catccccgag ccggcagcgt gcggggacag cccgggcacg
     1261 gggaaggtgg cacgggatcg ctttcctctg aacgcttctc gctgctcttt gagcctgcag
     1321 acacctgggg ggatacgggg aaaaggcctc cacggccact agtctttcgt cttcaagaat
     1381 tcctcgagtt tactccctat cagtgataga gaacgtatga agagtttact ccctatcagt
     1441 gatagagaac gtatgcagac tttactccct atcagtgata gagaacgtat aaggagttta
     1501 ctccctatca gtgatagaga acgtatgacc agtttactcc ctatcagtga tagagaacgt
     1561 atctacagtt tactccctat cagtgataga gaacgtatat ccagtttact ccctatcagt
     1621 gatagagaac gtataagctt taggcgtgta cggtgggcgc ctataaaagc agagctcgtt
     1681 tagtgaaccg tcagatcgcc tggagcaatt ccacaacact tttgtcttat accaactttc
     1741 cgtaccactt cctaccctcg taaaggtcta gagctagcca tgggccccga catcgtgatg
     1801 acccagagcc ccagcagcct gagcgccagc gtgggcgacc gcgtgaccat cacctgccgc
     1861 agcagcaccg gcgccgtgac caccagcaac tacgccagct gggtgcagga gaagcccggc
     1921 aagctgttca agggcctgat cggcggcacc aacaaccgcg cccccggcgt gcccagccgc
     1981 ttcagcggca gcctgatcgg cgacaaggcc accctgacca tcagcagcct gcagcccgag
     2041 gacttcgcca cctacttctg cgccctgtgg tacagcaacc actgggtgtt cggccagggc
     2101 accaaggtgg agctgaagcg tggaggcgga ggtagtggag gtggaggctc tggtggggga
     2161 ggaagcagtg gaggcggaag tgaggtgaag ctgctggaga gcggaggcgg actggtgcag
     2221 cccggcggat ctctgaagct gagctgcgcc gtgagcggct tcagcctgac cgactacggc
     2281 gtgaactggg tgcgccaggc ccccggacgc ggactggagt ggatcggtgt gatctggggc
     2341 gacggcatca ccgactacaa cagcgccctg aaggaccgct tcatcatcag caaggacaac
     2401 ggcaagaaca ccgtgtacct gcagatgagc aaggtgcgca gcgacgacac cgccctgtac
     2461 tactgcgtga ccggcctgtt cgactactgg ggccagggca ccctggtgac cgtgagcagc
     2521 tacccatacg atgttccaga ttacgctggt ggaggcggag gttctggggg aggaggtagt
     2581 ggcggtggtg gttcaggagg cggcggaagc ttggatccag gtggaggtgg aagcggtagc
     2641 aaaggagaag aacttttcac tggagttgtc ccaattcttg ttgaattaga tggtgatgtt
     2701 aatgggcaca aattttctgt ccgtggagag ggtgaaggtg atgctacaaa cggaaaactc
     2761 acccttaaat ttatttgcac tactggaaaa ctacctgttc cgtggccaac acttgtcact
     2821 actctgacct atggtgttca atgcttttcc cgttatccgg atcacatgaa acggcatgac
     2881 tttttcaaga gtgccatgcc cgaaggttat gtacaggaac gcactatatc tttcaaagat
     2941 gacgggacct acaagacgcg tgctgaagtc aagtttgaag gtgataccct tgttaatcgt
     3001 atcgagttaa agggtattga ttttaaagaa gatggaaaca ttcttggaca caaactcgag
     3061 tacaacttta actcacacaa tgtatacatc acggcagaca aacaaaagaa tggaatcaaa
     3121 gctaacttca aaattcgcca caacgttgaa gatggttccg ttcaactagc agaccattat
     3181 caacaaaata ctccaattgg cgatggccct gtccttttac cagacaacca ttacctgtcg
     3241 acacaatctg tcctttcgaa agatcccaac gaaaagcgtg accacatggt ccttcttgag
     3301 tttgtaactg ctgctgggat tacacatggc atggatgagc tctacaaagg tggaggtcgg
     3361 accggcccca agaaaaaacg caaggtggaa gatcctaaga aaaagcggaa agtggacggc
     3421 attggtagtg ggagcaacgg cagcagcgga tccggaggcg gtggtaccat gggccagact
     3481 gggaagaaat ctgagaaggg accggtttgt tggcggaagc gtgtaaaatc agagtacatg
     3541 agactgagac agctcaagag gttcagaaga gctgatgaag taaagactat gtttagttcc
     3601 aatcgtcaga aaattttgga aagaactgaa accttaaacc aagagtggaa gcagcggagg
     3661 atacagcctg tgcacatcat gacttctgtg agctcattgc gcgggactag ggagtgttca
     3721 gtcaccagtg acttggattt tccagcacaa gtcatcccgt taaagaccct gaatgcagtc
     3781 gcctcggtgc ctataatgta ctcttggtcg cccttacaac agaattttat ggtggaagac
     3841 gaaactgttt tacataacat tccttatatg ggggatgaag ttctggatca ggatggcact
     3901 ttcattgaag aactaataaa aaattatgat ggaaaagtgc atggtgacag agaatgtgga
     3961 tttataaatg atgaaatttt tgtggagttg gtaaatgctc ttggtcaata taatgatgat
     4021 gatgatgacg atgatggaga tgatccagat gaaagagaag aaaaacagaa agatctagag
     4081 gataatcgag atgataaaga aacttgccca cctcggaaat ttcctgctga taaaatattt
     4141 gaagccattt cctcaatgtt tccagataag ggcaccgcag aagaactgaa agaaaaatat
     4201 aaagaactca cggagcagca gctcccaggt gctctgcctc ctgaatgtac tccaaacatc
     4261 gatggaccaa atgccaaatc tgttcagagg gagcaaagct tgcattcatt tcatacgctc
     4321 ttctgtcgac gatgttttaa gtatgactgc ttcctacatc ccttccatgc aacacccaac
     4381 acatataaga ggaagaacac agaaacagct ttggacaaca agccttgtgg accacagtgt
     4441 taccagcatc tggagggagc taaggagttt gctgctgctc ttactgctga gcgtataaag
     4501 acaccaccta aacgcccagg gggccgcaga agaggaagac ttccgaataa cagtagcaga
     4561 cccagcaccc ccaccatcag tgtgctggag tcaaaggata cagacagtga cagagaagca
     4621 gggactgaaa ctgggggaga gaacaatgat aaagaagaag aagagaaaaa agatgagacg
     4681 tccagctcct ctgaagcaaa ttctcggtgt caaacaccaa taaagatgaa gccaaatatt
     4741 gaacctcctg agaatgtgga gtggagtggt gctgaagcct ccatgtttag agtcctcatt
     4801 ggtacttact acgataactt ttgtgccatt gctaggctaa ttgggaccaa aacatgtaga
     4861 caggtgtatg agtttagagt caaggagtcc agtatcatag cacctgttcc cactgaggat
     4921 gtagacactc ctccaagaaa gaagaaaagg aaacatcggt tgtgggctgc acactgcaga
     4981 aagatacaac tgaaaaagga cggctcctct aaccatgttt acaactatca accctgtgac
     5041 catccacggc agccttgtga cagttcgtgc ccttgtgtga tagcacaaaa tttttgtgaa
     5101 aagttttgtc aatgtagttc agagtgtcaa aaccgctttc ctggatgtcg gtgcaaagca
     5161 caatgcaaca ccaaacagtg tccatgctac ctggctgtcc gagagtgtga ccctgacctc
     5221 tgtctcacgt gtggagctgc tgaccattgg gacagtaaaa atgtatcctg taagaactgt
     5281 agcattcagc ggggctctaa aaagcactta ctgctggcac cgtctgatgt ggcaggctgg
     5341 ggcatcttta tcaaagatcc tgtacagaaa aatgaattca tctcagaata ctgtggggag
     5401 attatttctc aggatgaagc agacagaaga ggaaaagtgt atgacaaata catgtgcagc
     5461 tttctgttca acttgaacaa tgattttgtg gtggatgcaa cccgaaaggg caacaaaatt
     5521 cgttttgcta atcattcagt aaatccaaac tgctatgcaa aagttatgat ggttaatggt
     5581 gaccacagga taggcatctt tgctaagagg gctatccaga ctggtgaaga gttgtttttt
     5641 gattacagat acagccaggc tgatgccctg aagtatgtgg gcatcgaacg agaaatggaa
     5701 atcccttgac aaaagatcat aatcagccat atcacatctg tagaggtttt acttgcttta
     5761 aaaaacctcc cacacctccc cctgaacctg aaacataaaa tgaatgcaat tgttgttgtt
     5821 aacttgttta ttgcagctta taatggttac aaataaagca atagcatcac aaatttcaca
     5881 aataaagcat ttttttcact gcattctagt tgtggtttgt ccaaactcat caatgtatct
     5941 tatcatgtct catatggcct tctgcaggag cgtacagaac ccagggccct ggcacccgtg
     6001 cagaccctgg cccaccccac ctgggcgctc agtgcccaag agatgtccac acctaggatg
     6061 tcccgcggtg ggtggggggc ccgagagacg ggcaggccgg gggcaggcct ggccatgcgg
     6121 ggccgaaccg ggcactgccc agcgtgggcg cgggggccac ggcgcgcgcc cccagccccc
     6181 gggcccagca ccccaaggcg gccaacgcca aaactctccc tcctcctctt cctcaatctc
     6241 gctctcgctc tttttttttt tcgcaaaagg aggggagagg gggtaaaaaa atgctgcact
     6301 gtgcggcgaa gccggtgagt gagcggcgcg gggccaatca gcgtgcgccg ttccgaaagt
     6361 tgccttttat ggctcgagcg gccgcggcgg cgccctataa aacccagcgg cgcgacgcgc
     6421 caccaccgcc gagaccgcgt ccgccccgcg agcacagagc ctcgcctttg ccgatcctct
     6481 agagtcgaga tccgccgcca ccatgattga acaagatgga ttgcacgcag gttctccggc
     6541 cgcttgggtg gagaggctat tcggctatga ctgggcacaa cagacaatcg gctgctctga
     6601 tgccgccgtg ttccggctgt cagcgcaggg gcgcccggtt ctttttgtca agaccgacct
     6661 gtccggtgcc ctgaatgaac tgcaggacga ggcagcgcgg ctatcgtggc tggccacgac
     6721 gggcgttcct tgcgcagctg tgctcgacgt tgtcactgaa gcgggaaggg actggctgct
     6781 attgggcgaa gtgccggggc aggatctcct gtcatctcac cttgctcctg ccgagaaagt
     6841 atccatcatg gctgatgcaa tgcggcggct gcatacgctt gatccggcta cctgcccatt
     6901 cgaccaccaa gcgaaacatc gcatcgagcg agcacgtact cggatggaag ccggtcttgt
     6961 cgatcaggat gatctggacg aagagcatca ggggctcgcg ccagccgaac tgttcgccag
     7021 gctcaaggcg cgcatgcccg acggcgagga tctcgtcgtg acccatggcg atgcctgctt
     7081 gccgaatatc atggtggaaa atggccgctt ttctggattc atcgactgtg gccggctggg
     7141 tgtggcggac cgctatcagg acatagcgtt ggctacccgt gatattgctg aagagcttgg
     7201 cggcgaatgg gctgaccgct tcctcgtgct ttacggtatc gccgctcccg attcgcagcg
     7261 catcgccttc tatcgccttc ttgacgagtt cttctgagcg ggactctggg gttcgaaatg
     7321 accgaccaag cgacgcccaa cctgccatca cgagatttcg attccaccgc cgccttctat
     7381 gaaaggttgg gcttcggaat cgttttccgg gacgccggct ggatgatcct ccagcgcggg
     7441 gatctcatgc tggagttctt cgcccacccc ccggatctaa gctctagagt taacttgttt
     7501 attgcagctt ataatggtta caaataaagc aatagcatca caaatttcac aaataaagca
     7561 tttttttcac tgcattctag ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc
     7621 tggaattgac tcaaatgatg tcaattagtc tatcagaagc tatctggtct cccttccggg
     7681 ggacaagaca tccctgttta atatttaaac agcagtgttc ccaaactggg ttcttatatc
     7741 ccttgctctg gtcaaccagg ttgcagggtt tcctgtcctc acaggaacga agtccctaaa
     7801 gaaacagtgg cagccaggtt tagccccgga attgactgga ttcctttttt agggcccatt
     7861 ggtatggctt tttccccgta tccccccagg tgtctgcagg ctcaaagagc agcgagaagc
     7921 gttcagagga aagcgatccc gtgccacctt ccccgtgccc gggctgtccc cgcacgctgc
     7981 cggctcgggg atgcgggggg agcgccggac cggagcggag ccccgggcgg ctcgctgctg
     8041 ccccctagcg ggggagggac gtaattacat ccctgggggc tttggggggg ggctgtccct
     8101 gatatctata acaagaaaat atatatataa taagttatca cgtaagtaga acatgaaata
     8161 acaatataat tatcgtatga gttaaatctt aaaagtcacg taaaagataa tcatgcgtca
     8221 ttttgactca cgcggtcgtt atagttcaaa atcagtgaca cttaccgcat tgacaagcac
     8281 gcctcacggg agctccaagc ggcgactgag atgtcctaaa tgcacagcga cggattcgcg
     8341 ctatttagaa agagagagca atatttcaag aatgcatgcg tcaattttac gcagactatc
     8401 tttctagggt taatctagct gcatcaggat catatcgtcg ggtctttttt ccggctcagt
     8461 catcgcccaa gctggcgcta tctgggcatc ggggaggaag aagcccgtgc cttttcccgc
     8521 gaggttgaag cggcatggaa agagtttgcc gaggatgact gctgctgcat tgacgttgag
     8581 cgaaaacgca cgtttaccat gatgattcgg gaaggtgtgg ccatgcacgc ctttaacggt
     8641 gaactgttcg ttcaggccac ctgggatacc agttcgtcgc ggcttttccg gacacagttc
     8701 cggatggtca gcccgaagcg catcagcaac ccgaacaata ccggcgacag ccggaactgc
     8761 cgtgccggtg tgcagattaa tgacagcggt gcggcgctgg gatattacgt cagcgaggac
     8821 gggtatcctg gctggatgcc gcagaaatgg acatggatac cccgtgagtt acccggcggg
     8881 cgcgcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
     8941 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
     9001 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
     9061 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
     9121 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
     9181 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
     9241 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
     9301 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     9361 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     9421 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     9481 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     9541 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     9601 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     9661 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     9721 ctaactacgg ctacactaga aggacagtat ttggtatctg cgctctgctg aagccagtta
     9781 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     9841 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     9901 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     9961 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
    10021 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
    10081 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
    10141 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
    10201 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
    10261 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
    10321 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
    10381 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
    10441 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
    10501 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
    10561 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
    10621 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
    10681 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
    10741 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
    10801 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
    10861 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
    10921 t
//
