LOCUS       sequence-462119-       14396 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses dCas9-EWSR1_IDR-EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14396)
  AUTHORS   Li J, Liu S, Kim S, Goell J, Drum ZA, Flores JP, Ma AJ, Mahata B, 
            Escobar M, Raterink A, Ahn JH, Teran ER, Guerra-Resendez RS, Zhou Y,
            Yu B, Diehl MR, Wang GG, Gustavsson AK, Phanstiel DH, Hilton IB
  TITLE     Biomolecular condensation of human IDRs initiates endogenous 
            transcription via intrachromosomal looping or high-density promoter 
            localization.
  JOURNAL   Nucleic Acids Res. 2025 Feb 8;53(4):gkaf056. doi: 
            10.1093/nar/gkaf056.
  PUBMED    39933697
REFERENCE   2  (bases 1 to 14396)
  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: "Nucleic 
            Acids Res."; date: "2025-02-8"; volume: "53(4):gkaf056. doi"; pages:
            " 10.1093/nar/gkaf056"
COMMENT     Sequence Label: dCas9-EWSR1_IDR-EGFP
FEATURES             Location/Qualifiers
     source          1..14396
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(344..355)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             533..766
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(792..809)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    798..1022
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1068..1496
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1155..1174)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1365..1386
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1505..1525)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1510..1839
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1690..1825
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1752..1771
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        1887..1934
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1953..2327
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    2457..2578
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2494..2513)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2548..2567
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2627..2643)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2627..2643)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2640..2662)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2651..2667
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(2675..2681,2682..2699,2700..2705))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2720..2741
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2858..2875)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3029..3617)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3109..3128)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3788..4579,4580..4648))
                     /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     4411..4430
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4649..4753)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(4828..4847)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5019..5398
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5400..5598
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5553..5573
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             5616..5796
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    5843..5968
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    6461..6694
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             6879..6923
                     /codon_start=1
                     /product="antigenic peptide corresponding to amino acids 
                     655 to 669 of the HIV envelope protein gp41 (Lutje Hulsik 
                     et al., 2013)"
                     /label=gp41 peptide
                     /note="recognized by the 2H10 single-chain llama nanobody"
                     /translation="KNEQELLELDKWASL"
     misc_feature    7221..7338
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     misc_RNA        7470..7545
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        7607..7818
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      7837..7846
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             7849..7869
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(7894..7917,7918..7920,7921..10407,10408..10410,
                     10411..11994)
                     /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             7894..11994
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /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             11995..12042
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             join(12844..12846,12847..12849,12850..13560)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(12844..12846,12847..12849,12850..13560)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(13150..13169)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     13497..13518
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             13567..13590
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             13600..13656
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             13657..14253
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     14150..14170
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    join(14269..14396,1..461)
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(14322..14342)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
ORIGIN
        1 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
       61 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
      121 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
      181 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
      241 tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctgtgt
      301 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
      361 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
      421 ccctcagacg agtcggatct ccctttgggc cgcctccccg cgtcgacttt aagaccaatg
      481 acttacaagg cagctgtaga tcttagccac tttttaaaag aaaagggggg actggaaggg
      541 ctaattcact cccaacgaag acaagatctg ctttttgctt gtactgggtc tctctggtta
      601 gaccagatct gagcctggga gctctctggc taactaggga acccactgct taagcctcaa
      661 taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac
      721 tagagatccc tcagaccctt ttagtcagtg tggaaaatct ctagcagggc ccgtttaaac
      781 ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt gcccctcccc
      841 cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat aaaatgagga
      901 aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg tggggcagga
      961 cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg tgggctctat
     1021 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     1081 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     1141 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     1201 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     1261 cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat cgccctgata
     1321 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     1381 aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag ggattttgcc
     1441 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattaatt
     1501 ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc aggcagaagt
     1561 atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc aggctcccca
     1621 gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt cccgccccta
     1681 actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc ccatggctga
     1741 ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct attccagaag
     1801 tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg agcttgtata
     1861 tccattttcg gatctgatca gcacgtgttg acaattaatc atcggcatag tatatcggca
     1921 tagtataata cgacaaggtg aggaactaaa ccatggccaa gttgaccagt gccgttccgg
     1981 tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg gaccgaccgg ctcgggttct
     2041 cccgggactt cgtggaggac gacttcgccg gtgtggtccg ggacgacgtg accctgttca
     2101 tcagcgcggt ccaggaccag gtggtgccgg acaacaccct ggcctgggtg tgggtgcgcg
     2161 gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc cacgaacttc cgggacgcct
     2221 ccgggccggc catgaccgag atcggcgagc agccgtgggg gcgggagttc gccctgcgcg
     2281 acccggccgg caactgcgtg cacttcgtgg ccgaggagca ggactgacac gtgctacgag
     2341 atttcgattc caccgccgcc ttctatgaaa ggttgggctt cggaatcgtt ttccgggacg
     2401 ccggctggat gatcctccag cgcggggatc tcatgctgga gttcttcgcc caccccaact
     2461 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     2521 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     2581 atgtctgtat accgtcgacc tctagctaga gcttggcgta atcatggtca tagctgtttc
     2641 ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt
     2701 gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc
     2761 ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg
     2821 ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct
     2881 cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca
     2941 cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga
     3001 accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc
     3061 acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg
     3121 cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat
     3181 acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt
     3241 atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc
     3301 agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg
     3361 acttatcgcc actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg
     3421 gtgctacaga gttcttgaag tggtggccta actacggcta cactagaaga acagtatttg
     3481 gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg
     3541 gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca
     3601 gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga
     3661 acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga
     3721 tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt
     3781 ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt
     3841 catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat
     3901 ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag
     3961 caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct
     4021 ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt
     4081 tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg
     4141 cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca
     4201 aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt
     4261 tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat
     4321 gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac
     4381 cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa
     4441 aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt
     4501 tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt
     4561 tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa
     4621 gggcgacacg gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt
     4681 atcagggtta ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa
     4741 taggggttcc gcgcacattt ccccgaaaag tgccacctga cgtcgacgga tcgggagatc
     4801 tcccgatccc ctatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca
     4861 gtatctgctc cctgcttgtg tgttggaggt cgctgagtag tgcgcgagca aaatttaagc
     4921 tacaacaagg caaggcttga ccgacaattg catgaagaat ctgcttaggg ttaggcgttt
     4981 tgcgctgctt cgcgatgtac gggccagata tacgcgttga cattgattat tgactagtta
     5041 ttaatagtaa tcaattacgg ggtcattagt tcatagccca tatatggagt tccgcgttac
     5101 ataacttacg gtaaatggcc cgcctggctg accgcccaac gacccccgcc cattgacgtc
     5161 aataatgacg tatgttccca tagtaacgcc aatagggact ttccattgac gtcaatgggt
     5221 ggagtattta cggtaaactg cccacttggc agtacatcaa gtgtatcata tgccaagtac
     5281 gccccctatt gacgtcaatg acggtaaatg gcccgcctgg cattatgccc agtacatgac
     5341 cttatgggac tttcctactt ggcagtacat ctacgtatta gtcatcgcta ttaccatggt
     5401 gatgcggttt tggcagtaca tcaatgggcg tggatagcgg tttgactcac ggggatttcc
     5461 aagtctccac cccattgacg tcaatgggag tttgttttgg caccaaaatc aacgggactt
     5521 tccaaaatgt cgtaacaact ccgccccatt gacgcaaatg ggcggtaggc gtgtacggtg
     5581 ggaggtctat ataagcagcg cgttttgcct gtactgggtc tctctggtta gaccagatct
     5641 gagcctggga gctctctggc taactaggga acccactgct taagcctcaa taaagcttgc
     5701 cttgagtgct tcaagtagtg tgtgcccgtc tgttgtgtga ctctggtaac tagagatccc
     5761 tcagaccctt ttagtcagtg tggaaaatct ctagcagtgg cgcccgaaca gggacttgaa
     5821 agcgaaaggg aaaccagagg agctctctcg acgcaggact cggcttgctg aagcgcgcac
     5881 ggcaagaggc gaggggcggc gactggtgag tacgccaaaa attttgacta gcggaggcta
     5941 gaaggagaga gatgggtgcg agagcgtcag tattaagcgg gggagaatta gatcgcgatg
     6001 ggaaaaaatt cggttaaggc cagggggaaa gaaaaaatat aaattaaaac atatagtatg
     6061 ggcaagcagg gagctagaac gattcgcagt taatcctggc ctgttagaaa catcagaagg
     6121 ctgtagacaa atactgggac agctacaacc atcccttcag acaggatcag aagaacttag
     6181 atcattatat aatacagtag caaccctcta ttgtgtgcat caaaggatag agataaaaga
     6241 caccaaggaa gctttagaca agatagagga agagcaaaac aaaagtaaga ccaccgcaca
     6301 gcaagcggcc gctgatcttc agacctggag gaggagatat gagggacaat tggagaagtg
     6361 aattatataa atataaagta gtaaaaattg aaccattagg agtagcaccc accaaggcaa
     6421 agagaagagt ggtgcagaga gaaaaaagag cagtgggaat aggagctttg ttccttgggt
     6481 tcttgggagc agcaggaagc actatgggcg cagcgtcaat gacgctgacg gtacaggcca
     6541 gacaattatt gtctggtata gtgcagcagc agaacaattt gctgagggct attgaggcgc
     6601 aacagcatct gttgcaactc acagtctggg gcatcaagca gctccaggca agaatcctgg
     6661 ctgtggaaag atacctaaag gatcaacagc tcctggggat ttggggttgc tctggaaaac
     6721 tcatttgcac cactgctgtg ccttggaatg ctagttggag taataaatct ctggaacaga
     6781 tttggaatca cacgacctgg atggagtggg acagagaaat taacaattac acaagcttaa
     6841 tacactcctt aattgaagaa tcgcaaaacc agcaagaaaa gaatgaacaa gaattattgg
     6901 aattagataa atgggcaagt ttgtggaatt ggtttaacat aacaaattgg ctgtggtata
     6961 taaaattatt cataatgata gtaggaggct tggtaggttt aagaatagtt tttgctgtac
     7021 tttctatagt gaatagagtt aggcagggat attcaccatt atcgtttcag acccacctcc
     7081 caaccccgag gggacccgac aggcccgaag gaatagaaga agaaggtgga gagagagaca
     7141 gagacagatc cattcgatta gtgaacggat cggcactgcg tgcgccaatt ctgcagacaa
     7201 atggcagtat tcatccacaa ttttaaaaga aaagggggga ttggggggta cagtgcaggg
     7261 gaaagaatag tagacataat agcaacagac atacaaacta aagaattaca aaaacaaatt
     7321 acaaaaattc aaaattttcg ggtttattac agggacagca gagatccagt ttggttaatt
     7381 aaggtaccga tatcactagt gttccattaa ctgcgctaaa aataattttt aaatcttttt
     7441 tagcttcttg ctcttttttg tacgtctctg ttttagagct agaaatagca agttaaaata
     7501 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tgaattcgct
     7561 agctaggtct tgaaaggagt gggaattggc tccggtgccc gtcagtgggc agagcgcaca
     7621 tcgcccacag tccccgagaa gttgggggga ggggtcggca attgatccgg tgcctagaga
     7681 aggtggcgcg gggtaaactg ggaaagtgat gtcgtgtact ggctccgcct ttttcccgag
     7741 ggtgggggag aaccgtatat aagtgcagta gtcgccgtga acgttctttt tcgcaacggg
     7801 tttgccgcca gaacacagga ccggttctag agcgctgcca ccatggcccc aaagaagaag
     7861 cggaaggtcg gtatccacgg agtcccagca gccgacaaga agtacagcat cggcctggcc
     7921 atcggcacca actctgtggg ctgggccgtg atcaccgacg agtacaaggt gcccagcaag
     7981 aaattcaagg tgctgggcaa caccgaccgg cacagcatca agaagaacct gatcggagcc
     8041 ctgctgttcg acagcggcga aacagccgag gccacccggc tgaagagaac cgccagaaga
     8101 agatacacca gacggaagaa ccggatctgc tatctgcaag agatcttcag caacgagatg
     8161 gccaaggtgg acgacagctt cttccacaga ctggaagagt ccttcctggt ggaagaggat
     8221 aagaagcacg agcggcaccc catcttcggc aacatcgtgg acgaggtggc ctaccacgag
     8281 aagtacccca ccatctacca cctgagaaag aaactggtgg acagcaccga caaggccgac
     8341 ctgcggctga tctatctggc cctggcccac atgatcaagt tccggggcca cttcctgatc
     8401 gagggcgacc tgaaccccga caacagcgac gtggacaagc tgttcatcca gctggtgcag
     8461 acctacaacc agctgttcga ggaaaacccc atcaacgcca gcggcgtgga cgccaaggcc
     8521 atcctgtctg ccagactgag caagagcaga cggctggaaa atctgatcgc ccagctgccc
     8581 ggcgagaaga agaatggcct gttcggaaac ctgattgccc tgagcctggg cctgaccccc
     8641 aacttcaaga gcaacttcga cctggccgag gatgccaaac tgcagctgag caaggacacc
     8701 tacgacgacg acctggacaa cctgctggcc cagatcggcg accagtacgc cgacctgttt
     8761 ctggccgcca agaacctgtc cgacgccatc ctgctgagcg acatcctgag agtgaacacc
     8821 gagatcacca aggcccccct gagcgcctct atgatcaaga gatacgacga gcaccaccag
     8881 gacctgaccc tgctgaaagc tctcgtgcgg cagcagctgc ctgagaagta caaagagatt
     8941 ttcttcgacc agagcaagaa cggctacgcc ggctacattg acggcggagc cagccaggaa
     9001 gagttctaca agttcatcaa gcccatcctg gaaaagatgg acggcaccga ggaactgctc
     9061 gtgaagctga acagagagga cctgctgcgg aagcagcgga ccttcgacaa cggcagcatc
     9121 ccccaccaga tccacctggg agagctgcac gccattctgc ggcggcagga agatttttac
     9181 ccattcctga aggacaaccg ggaaaagatc gagaagatcc tgaccttccg catcccctac
     9241 tacgtgggcc ctctggccag gggaaacagc agattcgcct ggatgaccag aaagagcgag
     9301 gaaaccatca ccccctggaa cttcgaggaa gtggtggaca agggcgcttc cgcccagagc
     9361 ttcatcgagc ggatgaccaa cttcgataag aacctgccca acgagaaggt gctgcccaag
     9421 cacagcctgc tgtacgagta cttcaccgtg tataacgagc tgaccaaagt gaaatacgtg
     9481 accgagggaa tgagaaagcc cgccttcctg agcggcgagc agaaaaaggc catcgtggac
     9541 ctgctgttca agaccaaccg gaaagtgacc gtgaagcagc tgaaagagga ctacttcaag
     9601 aaaatcgagt gcttcgactc cgtggaaatc tccggcgtgg aagatcggtt caacgcctcc
     9661 ctgggcacat accacgatct gctgaaaatt atcaaggaca aggacttcct ggacaatgag
     9721 gaaaacgagg acattctgga agatatcgtg ctgaccctga cactgtttga ggacagagag
     9781 atgatcgagg aacggctgaa aacctatgcc cacctgttcg acgacaaagt gatgaagcag
     9841 ctgaagcggc ggagatacac cggctggggc aggctgagcc ggaagctgat caacggcatc
     9901 cgggacaagc agtccggcaa gacaatcctg gatttcctga agtccgacgg cttcgccaac
     9961 agaaacttca tgcagctgat ccacgacgac agcctgacct ttaaagagga catccagaaa
    10021 gcccaggtgt ccggccaggg cgatagcctg cacgagcaca ttgccaatct ggccggcagc
    10081 cccgccatta agaagggcat cctgcagaca gtgaaggtgg tggacgagct cgtgaaagtg
    10141 atgggccggc acaagcccga gaacatcgtg atcgaaatgg ccagagagaa ccagaccacc
    10201 cagaagggac agaagaacag ccgcgagaga atgaagcgga tcgaagaggg catcaaagag
    10261 ctgggcagcc agatcctgaa agaacacccc gtggaaaaca cccagctgca gaacgagaag
    10321 ctgtacctgt actacctgca gaatgggcgg gatatgtacg tggaccagga actggacatc
    10381 aaccggctgt ccgactacga tgtggacgcc atcgtgcctc agagctttct gaaggacgac
    10441 tccatcgaca acaaggtgct gaccagaagc gacaagaacc ggggcaagag cgacaacgtg
    10501 ccctccgaag aggtcgtgaa gaagatgaag aactactggc ggcagctgct gaacgccaag
    10561 ctgattaccc agagaaagtt cgacaatctg accaaggccg agagaggcgg cctgagcgaa
    10621 ctggataagg ccggcttcat caagagacag ctggtggaaa cccggcagat cacaaagcac
    10681 gtggcacaga tcctggactc ccggatgaac actaagtacg acgagaatga caagctgatc
    10741 cgggaagtga aagtgatcac cctgaagtcc aagctggtgt ccgatttccg gaaggatttc
    10801 cagttttaca aagtgcgcga gatcaacaac taccaccacg cccacgacgc ctacctgaac
    10861 gccgtcgtgg gaaccgccct gatcaaaaag taccctaagc tggaaagcga gttcgtgtac
    10921 ggcgactaca aggtgtacga cgtgcggaag atgatcgcca agagcgagca ggaaatcggc
    10981 aaggctaccg ccaagtactt cttctacagc aacatcatga actttttcaa gaccgagatt
    11041 accctggcca acggcgagat ccggaagcgg cctctgatcg agacaaacgg cgaaaccggg
    11101 gagatcgtgt gggataaggg ccgggatttt gccaccgtgc ggaaagtgct gagcatgccc
    11161 caagtgaata tcgtgaaaaa gaccgaggtg cagacaggcg gcttcagcaa agagtctatc
    11221 ctgcccaaga ggaacagcga taagctgatc gccagaaaga aggactggga ccctaagaag
    11281 tacggcggct tcgacagccc caccgtggcc tattctgtgc tggtggtggc caaagtggaa
    11341 aagggcaagt ccaagaaact gaagagtgtg aaagagctgc tggggatcac catcatggaa
    11401 agaagcagct tcgagaagaa tcccatcgac tttctggaag ccaagggcta caaagaagtg
    11461 aaaaaggacc tgatcatcaa gctgcctaag tactccctgt tcgagctgga aaacggccgg
    11521 aagagaatgc tggcctctgc cggcgaactg cagaagggaa acgaactggc cctgccctcc
    11581 aaatatgtga acttcctgta cctggccagc cactatgaga agctgaaggg ctcccccgag
    11641 gataatgagc agaaacagct gtttgtggaa cagcacaagc actacctgga cgagatcatc
    11701 gagcagatca gcgagttctc caagagagtg atcctggccg acgctaatct ggacaaagtg
    11761 ctgtccgcct acaacaagca ccgggataag cccatcagag agcaggccga gaatatcatc
    11821 cacctgttta ccctgaccaa tctgggagcc cctgccgcct tcaagtactt tgacaccacc
    11881 atcgaccgga agaggtacac cagcaccaaa gaggtgctgg acgccaccct gatccaccag
    11941 agcatcaccg gcctgtacga gacacggatc gacctgtctc agctgggagg cgacaagcga
    12001 cctgccgcca caaagaaggc tggacaggct aagaagaaga aaggatccgc gtccacggat
    12061 tacagtacct atagccaagc tgcagcgcag cagggctaca gtgcttacac cgcccagccc
    12121 actcaaggat atgcacagac cacccaggca tatgggcaac aaagctatgg aacctatgga
    12181 cagcccactg atgtcagcta tacccaggct cagaccactg caacctatgg gcagaccgcc
    12241 tatgcaactt cttatggaca gcctcccact ggttatacta ctccaactgc cccccaggca
    12301 tacagccagc ctgtccaggg gtatggcact ggtgcttatg ataccaccac tgctacagtc
    12361 accaccaccc aggcctccta tgcagctcag tctgcatatg gcactcagcc tgcttatcca
    12421 gcctatgggc agcagccagc agccactgca cctacaagac cgcaggatgg aaacaagccc
    12481 actgagacta gtcaacctca atctagcaca gggggttaca accagcccag cctaggatat
    12541 ggacagagta actacagtta tccccaggta cctgggagct accccatgca gccagtcact
    12601 gcacctccat cctaccctcc taccagctat tcctctacac agccgactag ttatgatcag
    12661 agcagttact ctcagcagaa cacctatggg caaccgagca gctatggaca gcagagtagc
    12721 tatggtcaac aaagcagcta tgggcagcag cctcccacta gttacccacc ccaaactgga
    12781 tcctacagcc aagctccaag tcaatatagc caacagagca gcagctacgg gcagcaggga
    12841 tccatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct ggtcgagctg
    12901 gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg cgatgccacc
    12961 tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt gccctggccc
    13021 accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc cgaccacatg
    13081 aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga gcgcaccatc
    13141 ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga gggcgacacc
    13201 ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa catcctgggg
    13261 cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga caagcagaag
    13321 aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag cgtgcagctc
    13381 gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct gcccgacaac
    13441 cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg cgatcacatg
    13501 gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga gctgtacaag
    13561 ggatccgatt acaaagacga tgacgataag ggatccggcg caacaaactt ctctctgctg
    13621 aaacaagccg gagatgtcga agagaatcct ggaccgaccg agtacaagcc cacggtgcgc
    13681 ctcgccaccc gcgacgacgt ccccagggcc gtacgcaccc tcgccgccgc gttcgccgac
    13741 taccccgcca cgcgccacac cgtcgatccg gaccgccaca tcgagcgggt caccgagctg
    13801 caagaactct tcctcacgcg cgtcgggctc gacatcggca aggtgtgggt cgcggacgac
    13861 ggcgccgcgg tggcggtctg gaccacgccg gagagcgtcg aagcgggggc ggtgttcgcc
    13921 gagatcggcc cgcgcatggc cgagttgagc ggttcccggc tggccgcgca gcaacagatg
    13981 gaaggcctcc tggcgccgca ccggcccaag gagcccgcgt ggttcctggc caccgtcgga
    14041 gtctcgcccg accaccaggg caagggtctg ggcagcgccg tcgtgctccc cggagtggag
    14101 gcggccgagc gcgccggggt gcccgccttc ctggagacct ccgcgccccg caacctcccc
    14161 ttctacgagc ggctcggctt caccgtcacc gccgacgtcg aggtgcccga aggaccgcgc
    14221 acctggtgca tgacccgcaa gcccggtgcc tgaacgcgtt aagtcgacaa tcaacctctg
    14281 gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta
    14341 tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat ggcttt
//
