LOCUS       sequence-257543-       13012 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Knock-out of human NONO.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13012)
  AUTHORS   Lahaye X, Gentili M, Silvin A, Conrad C, Picard L, Jouve M, Zueva E,
            Maurin M, Nadalin F, Knott GJ, Zhao B, Du F, Rio M, Amiel J, Fox AH,
            Li P, Etienne L, Bond CS, Colleaux L, Manel N
  TITLE     NONO Detects the Nuclear HIV Capsid to Promote cGAS-Mediated Innate 
            Immune Activation.
  JOURNAL   Cell. 2018 Oct 4;175(2):488-501.e22. doi: 
            10.1016/j.cell.2018.08.062. Epub 2018 Sep 27.
  PUBMED    30270045
REFERENCE   2  (bases 1 to 13012)
  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"; doi: 
            "10.1016/j.cell.2018.08.062"; journalName: "Cell"; date: "2018-10-4-
            4"; volume: "175"; issue: "2"; pages: "488-501.e22"
COMMENT     Sequence Label: pLentiCRISPRv2-NONO 4
FEATURES             Location/Qualifiers
     source          1..13012
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    186..303
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        354..602
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     354..374
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     525..544
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        623..698
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        760..971
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      990..999
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             996..5099
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             996..5099
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5100..5147
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5148..5171
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             5181..5237
                     /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             5238..5834
                     /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     5731..5751
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    5850..6438
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(5903..5923)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6321..6332)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             6510..6743
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(6769..6786)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6775..6999
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      7045..7473
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7132..7151)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7342..7363
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7482..7502)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        7487..7816
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      7667..7802
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     7729..7748
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        7864..7911
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             7930..8304
                     /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    8434..8555
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8471..8490)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8525..8544
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8604..8620)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8604..8620)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(8617..8639)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    8628..8644
                     /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(8652..8658,8659..8676,8677..8682))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8697..8718
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(8835..8852)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(9006..9594)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(9086..9105)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9765..10556,10557..10625))
                     /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     10388..10407
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10626..10730)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(10805..10824)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        10996..11375
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        11377..11575
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     11530..11550
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             11593..11773
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    11820..11945
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    12438..12671
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             12856..12900
                     /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"
ORIGIN
        1 gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac
       61 ccgacaggcc cgaaggaata gaagaagaag gtggagagag agacagagac agatccattc
      121 gattagtgaa cggatcggca ctgcgtgcgc caattctgca gacaaatggc agtattcatc
      181 cacaatttta aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac
      241 ataatagcaa cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat
      301 tttcgggttt attacaggga cagcagagat ccagtttggt taattaaggt accgagggcc
      361 tatttcccat gattccttca tatttgcata tacgatacaa ggctgttaga gagataatta
      421 gaattaattt gactgtaaac acaaagatat tagtacaaaa tacgtgacgt agaaagtaat
      481 aatttcttgg gtagtttgca gttttaaaat tatgttttaa aatggactat catatgctta
      541 ccgtaacttg aaagtatttc gatttcttgg ctttatatat cttgtggaaa ggacgaaaca
      601 ccgtatgtgt ccaacgaact gcgttttaga gctagaaata gcaagttaaa ataaggctag
      661 tccgttatca acttgaaaaa gtggcaccga gtcggtgctt ttttgaattc gctagctagg
      721 tcttgaaagg agtgggaatt ggctccggtg cccgtcagtg ggcagagcgc acatcgccca
      781 cagtccccga gaagttgggg ggaggggtcg gcaattgatc cggtgcctag agaaggtggc
      841 gcggggtaaa ctgggaaagt gatgtcgtgt actggctccg cctttttccc gagggtgggg
      901 gagaaccgta tataagtgca gtagtcgccg tgaacgttct ttttcgcaac gggtttgccg
      961 ccagaacaca ggaccggttc tagagcgctg ccaccatgga caagaagtac agcatcggcc
     1021 tggacatcgg caccaactct gtgggctggg ccgtgatcac cgacgagtac aaggtgccca
     1081 gcaagaaatt caaggtgctg ggcaacaccg accggcacag catcaagaag aacctgatcg
     1141 gagccctgct gttcgacagc ggcgaaacag ccgaggccac ccggctgaag agaaccgcca
     1201 gaagaagata caccagacgg aagaaccgga tctgctatct gcaagagatc ttcagcaacg
     1261 agatggccaa ggtggacgac agcttcttcc acagactgga agagtccttc ctggtggaag
     1321 aggataagaa gcacgagcgg caccccatct tcggcaacat cgtggacgag gtggcctacc
     1381 acgagaagta ccccaccatc taccacctga gaaagaaact ggtggacagc accgacaagg
     1441 ccgacctgcg gctgatctat ctggccctgg cccacatgat caagttccgg ggccacttcc
     1501 tgatcgaggg cgacctgaac cccgacaaca gcgacgtgga caagctgttc atccagctgg
     1561 tgcagaccta caaccagctg ttcgaggaaa accccatcaa cgccagcggc gtggacgcca
     1621 aggccatcct gtctgccaga ctgagcaaga gcagacggct ggaaaatctg atcgcccagc
     1681 tgcccggcga gaagaagaat ggcctgttcg gaaacctgat tgccctgagc ctgggcctga
     1741 cccccaactt caagagcaac ttcgacctgg ccgaggatgc caaactgcag ctgagcaagg
     1801 acacctacga cgacgacctg gacaacctgc tggcccagat cggcgaccag tacgccgacc
     1861 tgtttctggc cgccaagaac ctgtccgacg ccatcctgct gagcgacatc ctgagagtga
     1921 acaccgagat caccaaggcc cccctgagcg cctctatgat caagagatac gacgagcacc
     1981 accaggacct gaccctgctg aaagctctcg tgcggcagca gctgcctgag aagtacaaag
     2041 agattttctt cgaccagagc aagaacggct acgccggcta cattgacggc ggagccagcc
     2101 aggaagagtt ctacaagttc atcaagccca tcctggaaaa gatggacggc accgaggaac
     2161 tgctcgtgaa gctgaacaga gaggacctgc tgcggaagca gcggaccttc gacaacggca
     2221 gcatccccca ccagatccac ctgggagagc tgcacgccat tctgcggcgg caggaagatt
     2281 tttacccatt cctgaaggac aaccgggaaa agatcgagaa gatcctgacc ttccgcatcc
     2341 cctactacgt gggccctctg gccaggggaa acagcagatt cgcctggatg accagaaaga
     2401 gcgaggaaac catcaccccc tggaacttcg aggaagtggt ggacaagggc gcttccgccc
     2461 agagcttcat cgagcggatg accaacttcg ataagaacct gcccaacgag aaggtgctgc
     2521 ccaagcacag cctgctgtac gagtacttca ccgtgtataa cgagctgacc aaagtgaaat
     2581 acgtgaccga gggaatgaga aagcccgcct tcctgagcgg cgagcagaaa aaggccatcg
     2641 tggacctgct gttcaagacc aaccggaaag tgaccgtgaa gcagctgaaa gaggactact
     2701 tcaagaaaat cgagtgcttc gactccgtgg aaatctccgg cgtggaagat cggttcaacg
     2761 cctccctggg cacataccac gatctgctga aaattatcaa ggacaaggac ttcctggaca
     2821 atgaggaaaa cgaggacatt ctggaagata tcgtgctgac cctgacactg tttgaggaca
     2881 gagagatgat cgaggaacgg ctgaaaacct atgcccacct gttcgacgac aaagtgatga
     2941 agcagctgaa gcggcggaga tacaccggct ggggcaggct gagccggaag ctgatcaacg
     3001 gcatccggga caagcagtcc ggcaagacaa tcctggattt cctgaagtcc gacggcttcg
     3061 ccaacagaaa cttcatgcag ctgatccacg acgacagcct gacctttaaa gaggacatcc
     3121 agaaagccca ggtgtccggc cagggcgata gcctgcacga gcacattgcc aatctggccg
     3181 gcagccccgc cattaagaag ggcatcctgc agacagtgaa ggtggtggac gagctcgtga
     3241 aagtgatggg ccggcacaag cccgagaaca tcgtgatcga aatggccaga gagaaccaga
     3301 ccacccagaa gggacagaag aacagccgcg agagaatgaa gcggatcgaa gagggcatca
     3361 aagagctggg cagccagatc ctgaaagaac accccgtgga aaacacccag ctgcagaacg
     3421 agaagctgta cctgtactac ctgcagaatg ggcgggatat gtacgtggac caggaactgg
     3481 acatcaaccg gctgtccgac tacgatgtgg accatatcgt gcctcagagc tttctgaagg
     3541 acgactccat cgacaacaag gtgctgacca gaagcgacaa gaaccggggc aagagcgaca
     3601 acgtgccctc cgaagaggtc gtgaagaaga tgaagaacta ctggcggcag ctgctgaacg
     3661 ccaagctgat tacccagaga aagttcgaca atctgaccaa ggccgagaga ggcggcctga
     3721 gcgaactgga taaggccggc ttcatcaaga gacagctggt ggaaacccgg cagatcacaa
     3781 agcacgtggc acagatcctg gactcccgga tgaacactaa gtacgacgag aatgacaagc
     3841 tgatccggga agtgaaagtg atcaccctga agtccaagct ggtgtccgat ttccggaagg
     3901 atttccagtt ttacaaagtg cgcgagatca acaactacca ccacgcccac gacgcctacc
     3961 tgaacgccgt cgtgggaacc gccctgatca aaaagtaccc taagctggaa agcgagttcg
     4021 tgtacggcga ctacaaggtg tacgacgtgc ggaagatgat cgccaagagc gagcaggaaa
     4081 tcggcaaggc taccgccaag tacttcttct acagcaacat catgaacttt ttcaagaccg
     4141 agattaccct ggccaacggc gagatccgga agcggcctct gatcgagaca aacggcgaaa
     4201 ccggggagat cgtgtgggat aagggccggg attttgccac cgtgcggaaa gtgctgagca
     4261 tgccccaagt gaatatcgtg aaaaagaccg aggtgcagac aggcggcttc agcaaagagt
     4321 ctatcctgcc caagaggaac agcgataagc tgatcgccag aaagaaggac tgggacccta
     4381 agaagtacgg cggcttcgac agccccaccg tggcctattc tgtgctggtg gtggccaaag
     4441 tggaaaaggg caagtccaag aaactgaaga gtgtgaaaga gctgctgggg atcaccatca
     4501 tggaaagaag cagcttcgag aagaatccca tcgactttct ggaagccaag ggctacaaag
     4561 aagtgaaaaa ggacctgatc atcaagctgc ctaagtactc cctgttcgag ctggaaaacg
     4621 gccggaagag aatgctggcc tctgccggcg aactgcagaa gggaaacgaa ctggccctgc
     4681 cctccaaata tgtgaacttc ctgtacctgg ccagccacta tgagaagctg aagggctccc
     4741 ccgaggataa tgagcagaaa cagctgtttg tggaacagca caagcactac ctggacgaga
     4801 tcatcgagca gatcagcgag ttctccaaga gagtgatcct ggccgacgct aatctggaca
     4861 aagtgctgtc cgcctacaac aagcaccggg ataagcccat cagagagcag gccgagaata
     4921 tcatccacct gtttaccctg accaatctgg gagcccctgc cgccttcaag tactttgaca
     4981 ccaccatcga ccggaagagg tacaccagca ccaaagaggt gctggacgcc accctgatcc
     5041 accagagcat caccggcctg tacgagacac ggatcgacct gtctcagctg ggaggcgaca
     5101 agcgacctgc cgccacaaag aaggctggac aggctaagaa gaagaaagat tacaaagacg
     5161 atgacgataa gggatccggc gcaacaaact tctctctgct gaaacaagcc ggagatgtcg
     5221 aagagaatcc tggaccgacc gagtacaagc ccacggtgcg cctcgccacc cgcgacgacg
     5281 tccccagggc cgtacgcacc ctcgccgccg cgttcgccga ctaccccgcc acgcgccaca
     5341 ccgtcgatcc ggaccgccac atcgagcggg tcaccgagct gcaagaactc ttcctcacgc
     5401 gcgtcgggct cgacatcggc aaggtgtggg tcgcggacga cggcgccgcg gtggcggtct
     5461 ggaccacgcc ggagagcgtc gaagcggggg cggtgttcgc cgagatcggc ccgcgcatgg
     5521 ccgagttgag cggttcccgg ctggccgcgc agcaacagat ggaaggcctc ctggcgccgc
     5581 accggcccaa ggagcccgcg tggttcctgg ccaccgtcgg agtctcgccc gaccaccagg
     5641 gcaagggtct gggcagcgcc gtcgtgctcc ccggagtgga ggcggccgag cgcgccgggg
     5701 tgcccgcctt cctggagacc tccgcgcccc gcaacctccc cttctacgag cggctcggct
     5761 tcaccgtcac cgccgacgtc gaggtgcccg aaggaccgcg cacctggtgc atgacccgca
     5821 agcccggtgc ctgaacgcgt taagtcgaca atcaacctct ggattacaaa atttgtgaaa
     5881 gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac gctgctttaa
     5941 tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc ttgtataaat
     6001 cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt ggcgtggtgt
     6061 gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc tgtcagctcc
     6121 tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc gccgcctgcc
     6181 ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg gtgttgtcgg
     6241 ggaaatcatc gtcctttcct tggctgctcg cctgtgttgc cacctggatt ctgcgcggga
     6301 cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc cgcggcctgc
     6361 tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt cggatctccc
     6421 tttgggccgc ctccccgcgt cgactttaag accaatgact tacaaggcag ctgtagatct
     6481 tagccacttt ttaaaagaaa aggggggact ggaagggcta attcactccc aacgaagaca
     6541 agatctgctt tttgcttgta ctgggtctct ctggttagac cagatctgag cctgggagct
     6601 ctctggctaa ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca
     6661 agtagtgtgt gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta
     6721 gtcagtgtgg aaaatctcta gcagggcccg tttaaacccg ctgatcagcc tcgactgtgc
     6781 cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg accctggaag
     6841 gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat tgtctgagta
     6901 ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag gattgggaag
     6961 acaatagcag gcatgctggg gatgcggtgg gctctatggc ttctgaggcg gaaagaacca
     7021 gctggggctc tagggggtat ccccacgcgc cctgtagcgg cgcattaagc gcggcgggtg
     7081 tggtggttac gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg
     7141 ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg
     7201 ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt
     7261 agggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt
     7321 tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca ctcaacccta
     7381 tctcggtcta ttcttttgat ttataaggga ttttgccgat ttcggcctat tggttaaaaa
     7441 atgagctgat ttaacaaaaa tttaacgcga attaattctg tggaatgtgt gtcagttagg
     7501 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     7561 gtcagcaacc aggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     7621 gcatctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc cgcccctaac
     7681 tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta tttatgcaga
     7741 ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct tttttggagg
     7801 cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggat ctgatcagca
     7861 cgtgttgaca attaatcatc ggcatagtat atcggcatag tataatacga caaggtgagg
     7921 aactaaacca tggccaagtt gaccagtgcc gttccggtgc tcaccgcgcg cgacgtcgcc
     7981 ggagcggtcg agttctggac cgaccggctc gggttctccc gggacttcgt ggaggacgac
     8041 ttcgccggtg tggtccggga cgacgtgacc ctgttcatca gcgcggtcca ggaccaggtg
     8101 gtgccggaca acaccctggc ctgggtgtgg gtgcgcggcc tggacgagct gtacgccgag
     8161 tggtcggagg tcgtgtccac gaacttccgg gacgcctccg ggccggccat gaccgagatc
     8221 ggcgagcagc cgtgggggcg ggagttcgcc ctgcgcgacc cggccggcaa ctgcgtgcac
     8281 ttcgtggccg aggagcagga ctgacacgtg ctacgagatt tcgattccac cgccgccttc
     8341 tatgaaaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     8401 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     8461 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     8521 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     8581 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     8641 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     8701 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     8761 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     8821 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     8881 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     8941 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     9001 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     9061 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     9121 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     9181 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     9241 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     9301 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     9361 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     9421 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     9481 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     9541 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     9601 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     9661 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     9721 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     9781 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     9841 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     9901 ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     9961 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
    10021 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
    10081 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
    10141 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
    10201 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
    10261 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
    10321 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
    10381 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
    10441 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
    10501 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
    10561 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
    10621 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
    10681 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
    10741 cgaaaagtgc cacctgacgt cgacggatcg ggagatctcc cgatccccta tggtgcactc
    10801 tcagtacaat ctgctctgat gccgcatagt taagccagta tctgctccct gcttgtgtgt
    10861 tggaggtcgc tgagtagtgc gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg
    10921 acaattgcat gaagaatctg cttagggtta ggcgttttgc gctgcttcgc gatgtacggg
    10981 ccagatatac gcgttgacat tgattattga ctagttatta atagtaatca attacggggt
    11041 cattagttca tagcccatat atggagttcc gcgttacata acttacggta aatggcccgc
    11101 ctggctgacc gcccaacgac ccccgcccat tgacgtcaat aatgacgtat gttcccatag
    11161 taacgccaat agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc
    11221 acttggcagt acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg
    11281 gtaaatggcc cgcctggcat tatgcccagt acatgacctt atgggacttt cctacttggc
    11341 agtacatcta cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca
    11401 atgggcgtgg atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca
    11461 atgggagttt gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg
    11521 ccccattgac gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagcgcgt
    11581 tttgcctgta ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa
    11641 ctagggaacc cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt
    11701 gcccgtctgt tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg
    11761 aaaatctcta gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc
    11821 tctctcgacg caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac
    11881 tggtgagtac gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga
    11941 gcgtcagtat taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag
    12001 ggggaaagaa aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat
    12061 tcgcagttaa tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc
    12121 tacaaccatc ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa
    12181 ccctctattg tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga
    12241 tagaggaaga gcaaaacaaa agtaagacca ccgcacagca agcggccgct gatcttcaga
    12301 cctggaggag gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta
    12361 aaaattgaac cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa
    12421 aaaagagcag tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact
    12481 atgggcgcag cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg
    12541 cagcagcaga acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca
    12601 gtctggggca tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat
    12661 caacagctcc tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct
    12721 tggaatgcta gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg
    12781 gagtgggaca gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg
    12841 caaaaccagc aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg
    12901 tggaattggt ttaacataac aaattggctg tggtatataa aattattcat aatgatagta
    12961 ggaggcttgg taggtttaag aatagttttt gctgtacttt ctatagtgaa ta
//
