LOCUS       Exported               15948 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Targeted CpG methylation.
ACCESSION   .
VERSION     .
KEYWORDS    pLV hUbC-dCas9-MQ1(Q147L)-EGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 15948)
  AUTHORS   Lei Y, Zhang X, Su J, Jeong M, Gundry MC, Huang YH, Zhou Y, Li W, 
            Goodell MA
  TITLE     Targeted DNA methylation in vivo using an engineered dCas9-MQ1 
            fusion protein.
  JOURNAL   Nat Commun. 2017 Jul 11;8:16026. doi: 10.1038/ncomms16026.
  PUBMED    28695892
REFERENCE   2  (bases 1 to 15948)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..15948
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        49..1260
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     1097..1116
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      1289..1298
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1298..1363
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1370..1390
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1400..5503
                     /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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             5516..5536
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6707..6760
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             6767..7480
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             6767..7480
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(6809..6830)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(7070..7089)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     7417..7438
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    7496..7529
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    7554..8142
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7607..7627)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8025..8036)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(8145..8161)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8146..8162)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             8667..8847
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(8873..8890)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8879..9103
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9149..9577
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9236..9255)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9446..9467
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9586..9606)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9591..9920
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9771..9906
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9833..9852
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9968..10015
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10034..10408
                     /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    10538..10659
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10575..10594)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10629..10648
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10708..10724)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    10732..10748
                     /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(10756..10786)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10801..10822
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10939..10956)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11110..11698)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11190..11209)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(11869..12729)
                     /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     12492..12511
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12730..12834)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(12909..12928)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        13100..13479
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        13481..13679
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     13634..13654
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             13697..13877
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    13924..14049
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    14542..14775
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             14960..15004
                     /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    15302..15419
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    15464..15497
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     CDS             complement(15484..15810)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGATTVSY
                     HNFV"
     primer_bind     15660..15682
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     15674..15691
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     15675..15691
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     15701..15720
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        15701..15719
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(15767..15787)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(15767..15785)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(15806..15822)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    15830..15846
                     /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(15854..15884)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    15899..15920
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
ORIGIN
        1 actgcatgac gtctccacaa ttaattaacc cgtgtcggct ccagatctgg cctccgcgcc
       61 gggttttggc gcctcccgcg ggcgcccccc tcctcacggc gagcgctgcc acgtcagacg
      121 aagggcgcag cgagcgtcct gatccttccg cccggacgct caggacagcg gcccgctgct
      181 cataagactc ggccttagaa ccccagtatc agcagaagga cattttagga cgggacttgg
      241 gtgactctag ggcactggtt ttctttccag agagcggaac aggcgaggaa aagtagtccc
      301 ttctcggcga ttctgcggag ggatctccgt ggggcggtga acgccgatga ttatataagg
      361 acgcgccggg tgtggcacag ctagttccgt cgcagccggg atttgggtcg cggttcttgt
      421 ttgtggatcg ctgtgatcgt cacttggtga gtagcgggct gctgggctgg ccggggcttt
      481 cgtggccgcc gggccgctcg gtgggacgga agcgtgtgga gagaccgcca agggctgtag
      541 tctgggtccg cgagcaaggt tgccctgaac tgggggttgg ggggagcgca gcaaaatggc
      601 ggctgttccc gagtcttgaa tggaagacgc ttgtgaggcg ggctgtgagg tcgttgaaac
      661 aaggtggggg gcatggtggg cggcaagaac ccaaggtctt gaggccttcg ctaatgcggg
      721 aaagctctta ttcgggtgag atgggctggg gcaccatctg gggaccctga cgtgaagttt
      781 gtcactgact ggagaactcg gtttgtcgtc tgttgcgggg gcggcagtta tggcggtgcc
      841 gttgggcagt gcacccgtac ctttgggagc gcgcgccctc gtcgtgtcgt gacgtcaccc
      901 gttctgttgg cttataatgc agggtggggc cacctgccgg taggtgtgcg gtaggctttt
      961 ctccgtcgca ggacgcaggg ttcgggccta gggtaggctc tcctgaatcg acaggcgccg
     1021 gacctctggt gaggggaggg ataagtgagg cgtcagtttc tttggtcggt tttatgtacc
     1081 tatcttctta agtagctgaa gctccggttt tgaactatgc gctcggggtt ggcgagtgtg
     1141 ttttgtgaag ttttttaggc accttttgaa atgtaatcat ttgggtcaat atgtaatttt
     1201 cagtgttaga ctagtaaatt gtccgctaaa ttctggccgt ttttggcttt tttgttagac
     1261 gaagcttggg ctgcaggtcg actctagagc caccatggac tacaaagacc atgacggtga
     1321 ttataaagat catgacatcg attacaagga tgacgatgac aagatggccc ccaagaagaa
     1381 gaggaaggtg ggccgcggaa tggacaagaa gtactccatt gggctcgcca tcggcacaaa
     1441 cagcgtcggc tgggccgtca ttacggacga gtacaaggtg ccgagcaaaa aattcaaagt
     1501 tctgggcaat accgatcgcc acagcataaa gaagaacctc attggcgccc tcctgttcga
     1561 ctccggggaa accgccgaag ccacgcggct caaaagaaca gcacggcgca gatatacccg
     1621 cagaaagaat cggatctgct acctgcagga gatctttagt aatgagatgg ctaaggtgga
     1681 tgactctttc ttccataggc tggaggagtc ctttttggtg gaggaggata aaaagcacga
     1741 gcgccaccca atctttggca atatcgtgga cgaggtggcg taccatgaaa agtacccaac
     1801 catatatcat ctgaggaaga agcttgtaga cagtactgat aaggctgact tgcggttgat
     1861 ctatctcgcg ctggcgcata tgatcaaatt tcggggacac ttcctcatcg agggggacct
     1921 gaacccagac aacagcgatg tcgacaaact ctttatccaa ctggttcaga cttacaatca
     1981 gcttttcgaa gagaacccga tcaacgcatc cggagttgac gccaaagcaa tcctgagcgc
     2041 taggctgtcc aaatcccggc ggctcgaaaa cctcatcgca cagctccctg gggagaagaa
     2101 gaacggcctg tttggtaatc ttatcgccct gtcactcggg ctgaccccca actttaaatc
     2161 taacttcgac ctggccgaag atgccaagct tcaactgagc aaagacacct acgatgatga
     2221 tctcgacaat ctgctggccc agatcggcga ccagtacgca gacctttttt tggcggcaaa
     2281 gaacctgtca gacgccattc tgctgagtga tattctgcga gtgaacacgg agatcaccaa
     2341 agctccgctg agcgctagta tgatcaagcg ctatgatgag caccaccaag acttgacttt
     2401 gctgaaggcc cttgtcagac agcaactgcc tgagaagtac aaggaaattt tcttcgatca
     2461 gtctaaaaat ggctacgccg gatacattga cggcggagca agccaggagg aattttacaa
     2521 atttattaag cccatcttgg aaaaaatgga cggcaccgag gagctgctgg taaagcttaa
     2581 cagagaagat ctgttgcgca aacagcgcac tttcgacaat ggaagcatcc cccaccagat
     2641 tcacctgggc gaactgcacg ctatcctcag gcggcaagag gatttctacc cctttttgaa
     2701 agataacagg gaaaagattg agaaaatcct cacatttcgg ataccctact atgtaggccc
     2761 cctcgcccgg ggaaattcca gattcgcgtg gatgactcgc aaatcagaag agaccatcac
     2821 tccctggaac ttcgaggaag tcgtggataa gggggcctct gcccagtcct tcatcgaaag
     2881 gatgactaac tttgataaaa atctgcctaa cgaaaaggtg cttcctaaac actctctgct
     2941 gtacgagtac ttcacagttt ataacgagct caccaaggtc aaatacgtca cagaagggat
     3001 gagaaagcca gcattcctgt ctggagagca gaagaaagct atcgtggacc tcctcttcaa
     3061 gacgaaccgg aaagttaccg tgaaacagct caaagaagac tatttcaaaa agattgaatg
     3121 tttcgactct gttgaaatca gcggagtgga ggatcgcttc aacgcatccc tgggaacgta
     3181 tcacgatctc ctgaaaatca ttaaagacaa ggacttcctg gacaatgagg agaacgagga
     3241 cattcttgag gacattgtcc tcacccttac gttgtttgaa gatagggaga tgattgaaga
     3301 acgcttgaaa acttacgctc atctcttcga cgacaaagtc atgaaacagc tcaagaggcg
     3361 ccgatataca ggatgggggc ggctgtcaag aaaactgatc aatgggatcc gagacaagca
     3421 gagtggaaag acaatcctgg attttcttaa gtccgatgga tttgccaacc ggaacttcat
     3481 gcagttgatc catgatgact ctctcacctt taaggaggac atccagaaag cacaagtttc
     3541 tggccagggg gacagtcttc acgagcacat cgctaatctt gcaggtagcc cagctatcaa
     3601 aaagggaata ctgcagaccg ttaaggtcgt ggatgaactc gtcaaagtaa tgggaaggca
     3661 taagcccgag aatatcgtta tcgagatggc ccgagagaac caaactaccc agaagggaca
     3721 gaagaacagt agggaaagga tgaagaggat tgaagagggt ataaaagaac tggggtccca
     3781 aatccttaag gaacacccag ttgaaaacac ccagcttcag aatgagaagc tctacctgta
     3841 ctacctgcag aacggcaggg acatgtacgt ggatcaggaa ctggacatca atcggctctc
     3901 cgactacgac gtggatgcca tcgtgcccca gtcttttctc aaagatgatt ctattgataa
     3961 taaagtgttg acaagatccg ataaaaatag agggaagagt gataacgtcc cctcagaaga
     4021 agttgtcaag aaaatgaaaa attattggcg gcagctgctg aacgccaaac tgatcacaca
     4081 acggaagttc gataatctga ctaaggctga acgaggtggc ctgtctgagt tggataaagc
     4141 cggcttcatc aaaaggcagc ttgttgagac acgccagatc accaagcacg tggcccaaat
     4201 tctcgattca cgcatgaaca ccaagtacga tgaaaatgac aaactgattc gagaggtgaa
     4261 agttattact ctgaagtcta agctggtctc agatttcaga aaggactttc agttttataa
     4321 ggtgagagag atcaacaatt accaccatgc gcatgatgcc tacctgaatg cagtggtagg
     4381 cactgcactt atcaaaaaat atcccaagct tgaatctgaa tttgtttacg gagactataa
     4441 agtgtacgat gttaggaaaa tgatcgcaaa gtctgagcag gaaataggca aggccaccgc
     4501 taagtacttc ttttacagca atattatgaa ttttttcaag accgagatta cactggccaa
     4561 tggagagatt cggaagcgac cacttatcga aacaaacgga gaaacaggag aaatcgtgtg
     4621 ggacaagggt agggatttcg cgacagtccg gaaggtcctg tccatgccgc aggtgaacat
     4681 cgttaaaaag accgaagtac agaccggagg cttctccaag gaaagtatcc tcccgaaaag
     4741 gaacagcgac aagctgatcg cacgcaaaaa agattgggac cccaagaaat acggcggatt
     4801 cgattctcct acagtcgctt acagtgtact ggttgtggcc aaagtggaga aagggaagtc
     4861 taaaaaactc aaaagcgtca aggaactgct gggcatcaca atcatggagc gatcaagctt
     4921 cgaaaaaaac cccatcgact ttctcgaggc gaaaggatat aaagaggtca aaaaagacct
     4981 catcattaag cttcccaagt actctctctt tgagcttgaa aacggccgga aacgaatgct
     5041 cgctagtgcg ggcgagctgc agaaaggtaa cgagctggca ctgccctcta aatacgttaa
     5101 tttcttgtat ctggccagcc actatgaaaa gctcaaaggg tctcccgaag ataatgagca
     5161 gaagcagctg ttcgtggaac aacacaaaca ctaccttgat gagatcatcg agcaaataag
     5221 cgaattctcc aaaagagtga tcctcgccga cgctaacctc gataaggtgc tttctgctta
     5281 caataagcac agggataagc ccatcaggga gcaggcagaa aacattatcc acttgtttac
     5341 tctgaccaac ttgggcgcgc ctgcagcctt caagtacttc gacaccacca tagacagaaa
     5401 gcggtacacc tctacaaagg aggtcctgga cgccacactg attcatcagt caattacggg
     5461 gctctatgaa acaagaatcg acctctctca gctcggtgga gacagcaggg ctgaccccaa
     5521 gaagaagagg aaggtggcta gggacagcaa agtggagaac aaaacaaaga agctgagagt
     5581 gttcgaagcc ttcgccggca ttggcgccca gagaaaggcc ctggagaaag tgaggaagga
     5641 cgagtacgag atcgtgggac tggccgagtg gtatgtgccc gccatcgtca tgtaccaggc
     5701 catccataac aacttccaca ccaagctcga gtacaagtcc gtcagcagag aggagatgat
     5761 cgactacctc gagaacaaga ccctgtcctg gaacagcaag aaccccgtca gcaacggata
     5821 ctggaagagg aagaaggatg acgagctgaa gatcatctac aacgccatca agctgtccga
     5881 aaaggagggc aacattttcg acatcaggga cctctacaag aggacactga agaacatcga
     5941 cctgctcacc tacagcttcc cttgccagga cctgagccag ctaggcatcc agaagggcat
     6001 gaagagggga agcggcacca gatccggcct gctctgggag atcgaaagag ccctggactc
     6061 caccgagaag aacgacctgc ctaagtacct cctcatggag aacgtgggag ccctgctgca
     6121 caagaagaac gaggaggagc tgaaccaatg gaagcagaag ctggagagcc tgggctacca
     6181 gaacagcatc gaagtcctca atgctgccga tttcggatcc agccaggcca ggaggagagt
     6241 gttcatgatc tccaccctca atgagttcgt ggaactgcct aagggcgaca agaagcccaa
     6301 gagcatcaaa aaggtgctga acaagatcgt gagcgagaag gacatcctca acaacctgct
     6361 gaaatacaac ctcaccgaat tcaagaagac caagtccaac atcaacaagg ccagcctgat
     6421 cggctactcc aagttcaact ccgagggcta tgtgtacgac cccgagttca caggccccac
     6481 actgacagct agcggcgcca actccaggat caagatcaag gacggcagca acatcaggaa
     6541 gatgaacagc gacgagacct tcctgtacat cggctttgac agccaggacg gcaagagagt
     6601 gaacgaaatc gagttcctga ccgagaacca gaagatcttc gtgtgtggca acagcatcag
     6661 cgtggaggtg ctggaggcca tcattgacaa gatcggcggc cctagcgagg gcagaggaag
     6721 tcttctaaca tgcggtgacg tggaggagaa tcccggccct ggtaccgtga gcaagggcga
     6781 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     6841 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     6901 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     6961 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     7021 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     7081 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     7141 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     7201 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
     7261 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     7321 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     7381 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     7441 cgccgccggg atcactctcg gcatggacga gctgtacaag accggttgat aatagataac
     7501 ttcgtatagc atacattata cgaagttatg aattcgatat caagcttatc gataatcaac
     7561 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     7621 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     7681 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     7741 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     7801 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     7861 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     7921 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg
     7981 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     8041 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     8101 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgtcgacctc
     8161 gagacctaga aaaacatgga gcaatcacaa gtagcaatac agcagctacc aatgctgatt
     8221 gtgcctggct agaagcacaa gaggaggagg aggtgggttt tccagtcaca cctcaggtac
     8281 ctttaagacc aatgacttac aaggcagctg tagatcttag ccacttttta aaagaaaagg
     8341 ggggactgga agggctaatt cactcccaac gaagacaaga tatccttgat ctgtggatct
     8401 accacacaca aggctacttc cctgattggc agaactacac accagggcca gggatcagat
     8461 atccactgac ctttggatgg tgctacaagc tagtaccagt tgagcaagag aaggtagaag
     8521 aagccaatga aggagagaac acccgcttgt tacaccctgt gagcctgcat gggatggatg
     8581 acccggagag agaagtatta gagtggaggt ttgacagccg cctagcattt catcacatgg
     8641 cccgagagct gcatccggac tgtactgggt ctctctggtt agaccagatc tgagcctggg
     8701 agctctctgg ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc
     8761 ttcaagtagt gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct
     8821 tttagtcagt gtggaaaatc tctagcaggg cccgtttaaa cccgctgatc agcctcgact
     8881 gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc cttgaccctg
     8941 gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc gcattgtctg
     9001 agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg ggaggattgg
     9061 gaagacaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga ggcggaaaga
     9121 accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt aagcgcggcg
     9181 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     9241 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     9301 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     9361 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     9421 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     9481 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     9541 aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt
     9601 tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc atgcatctca
     9661 attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga agtatgcaaa
     9721 gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc atcccgcccc
     9781 taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt tttatttatg
     9841 cagaggccga ggccgcctct gcctctgagc tattccagaa gtagtgagga ggcttttttg
     9901 gaggcctagg cttttgcaaa aagctcccgg gagcttgtat atccattttc ggatctgatc
     9961 agcacgtgtt gacaattaat catcggcata gtatatcggc atagtataat acgacaaggt
    10021 gaggaactaa accatggcca agttgaccag tgccgttccg gtgctcaccg cgcgcgacgt
    10081 cgccggagcg gtcgagttct ggaccgaccg gctcgggttc tcccgggact tcgtggagga
    10141 cgacttcgcc ggtgtggtcc gggacgacgt gaccctgttc atcagcgcgg tccaggacca
    10201 ggtggtgccg gacaacaccc tggcctgggt gtgggtgcgc ggcctggacg agctgtacgc
    10261 cgagtggtcg gaggtcgtgt ccacgaactt ccgggacgcc tccgggccgg ccatgaccga
    10321 gatcggcgag cagccgtggg ggcgggagtt cgccctgcgc gacccggccg gcaactgcgt
    10381 gcacttcgtg gccgaggagc aggactgaca cgtgctacga gatttcgatt ccaccgccgc
    10441 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
    10501 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
    10561 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
    10621 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
    10681 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
    10741 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
    10801 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
    10861 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
    10921 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
    10981 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
    11041 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
    11101 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
    11161 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
    11221 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
    11281 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
    11341 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
    11401 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
    11461 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
    11521 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
    11581 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
    11641 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
    11701 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
    11761 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
    11821 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
    11881 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
    11941 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
    12001 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
    12061 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
    12121 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
    12181 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
    12241 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
    12301 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
    12361 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
    12421 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
    12481 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
    12541 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
    12601 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
    12661 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
    12721 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
    12781 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
    12841 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc
    12901 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt
    12961 gtgttggagg tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg
    13021 accgacaatt gcatgaagaa tctgcttagg gttaggcgtt ttgcgctgct tcgcgatgta
    13081 cgggccagat atacgcgttg acattgatta ttgactagtt attaatagta atcaattacg
    13141 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
    13201 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
    13261 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
    13321 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
    13381 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
    13441 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
    13501 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
    13561 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
    13621 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcagc
    13681 gcgttttgcc tgtactgggt ctctctggtt agaccagatc tgagcctggg agctctctgg
    13741 ctaactaggg aacccactgc ttaagcctca ataaagcttg ccttgagtgc ttcaagtagt
    13801 gtgtgcccgt ctgttgtgtg actctggtaa ctagagatcc ctcagaccct tttagtcagt
    13861 gtggaaaatc tctagcagtg gcgcccgaac agggacttga aagcgaaagg gaaaccagag
    13921 gagctctctc gacgcaggac tcggcttgct gaagcgcgca cggcaagagg cgaggggcgg
    13981 cgactggtga gtacgccaaa aattttgact agcggaggct agaaggagag agatgggtgc
    14041 gagagcgtca gtattaagcg ggggagaatt agatcgcgat gggaaaaaat tcggttaagg
    14101 ccagggggaa agaaaaaata taaattaaaa catatagtat gggcaagcag ggagctagaa
    14161 cgattcgcag ttaatcctgg cctgttagaa acatcagaag gctgtagaca aatactggga
    14221 cagctacaac catcccttca gacaggatca gaagaactta gatcattata taatacagta
    14281 gcaaccctct attgtgtgca tcaaaggata gagataaaag acaccaagga agctttagac
    14341 aagatagagg aagagcaaaa caaaagtaag accaccgcac agcaagcggc cgctgatctt
    14401 cagacctgga ggaggagata tgagggacaa ttggagaagt gaattatata aatataaagt
    14461 agtaaaaatt gaaccattag gagtagcacc caccaaggca aagagaagag tggtgcagag
    14521 agaaaaaaga gcagtgggaa taggagcttt gttccttggg ttcttgggag cagcaggaag
    14581 cactatgggc gcagcgtcaa tgacgctgac ggtacaggcc agacaattat tgtctggtat
    14641 agtgcagcag cagaacaatt tgctgagggc tattgaggcg caacagcatc tgttgcaact
    14701 cacagtctgg ggcatcaagc agctccaggc aagaatcctg gctgtggaaa gatacctaaa
    14761 ggatcaacag ctcctgggga tttggggttg ctctggaaaa ctcatttgca ccactgctgt
    14821 gccttggaat gctagttgga gtaataaatc tctggaacag atttggaatc acacgacctg
    14881 gatggagtgg gacagagaaa ttaacaatta cacaagctta atacactcct taattgaaga
    14941 atcgcaaaac cagcaagaaa agaatgaaca agaattattg gaattagata aatgggcaag
    15001 tttgtggaat tggtttaaca taacaaattg gctgtggtat ataaaattat tcataatgat
    15061 agtaggaggc ttggtaggtt taagaatagt ttttgctgta ctttctatag tgaatagagt
    15121 taggcaggga tattcaccat tatcgtttca gacccacctc ccaaccccga ggggacccga
    15181 caggcccgaa ggaatagaag aagaaggtgg agagagagac agagacagat ccattcgatt
    15241 agtgaacgga tcggcactgc gtgcgccaat tctgcagaca aatggcagta ttcatccaca
    15301 attttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata gtagacataa
    15361 tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaaatt caaaattttc
    15421 gggtttatta cagggacagc agagatccag tttggttaat taaataactt cgtatagcat
    15481 acattatacg aagttatgat aagagacggt ggtggcgccg ctacagggcg cgtcccattc
    15541 gccattcagg ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg
    15601 ccagctggcg aaagggggat gtgctgcaag gcgattaagt tgggtaacgc cagggttttc
    15661 ccagtcacga cgttgtaaaa cgacggccag tgagcgcgcg taatacgact cactataggg
    15721 cgaattgggt accgggcccc ccctcgaggt cctccagctt ttgttccctt tagtgagggt
    15781 taattgcgcg cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc
    15841 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
    15901 gagtgagcta actcacatta attgcgttgc gctcaatgcc cgctttcc
//
