LOCUS       Exported               14833 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses dead Cas9 (dCas9) fused to DNMT3A catalytic domain (CD) 
            linked to IRES-mCherry under the control of TetOp and a minimal CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    TetO-dCas9-D3A
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14833)
  AUTHORS   McDonald JI, Celik H, Rois LE, Fishberger G, Fowler T, Rees R, 
            Kramer A, Martens A, Edwards JR, Challen GA
  TITLE     Reprogrammable CRISPR/Cas9-based system for inducing site-specific 
            DNA methylation.
  JOURNAL   Biol Open. 2016 May 11. pii: bio.019067. doi: 10.1242/bio.019067.
  PUBMED    27170255
REFERENCE   2  (bases 1 to 14833)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..14833
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     enhancer        74..453
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        455..653
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     608..628
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             671..851
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    898..1023
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1516..1749
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1934..1978
                     /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    2276..2393
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     protein_bind    2450..2720
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        2753..2791
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2788..2812
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     regulatory      2912..2921
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2918..7021
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=Cas9m4
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the mutations D10A, D839A, 
                     H840A, and N863A (Mali et al., 2013)"
                     /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEIAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     AIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             2918..7021
                     /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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEIAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     AIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7034..7054
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7064..7087
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             7106..8038
                     /codon_start=1
                     /product="catalytic domain of DNA methyltransferase 3A"
                     /label=DNMT3A catalytic domain
                     /translation="PSRLQMFFANNHDQEFDPPKVYPPVPAEKRKPIRVLSLFDGIATG
                     LLVLKDLGIQVDRYIASEVCEDSITVGMVRHQGKIMYVGDVRSVTQKHIQEWGPFDLVI
                     GGSPCNDLSIVNPARKGLYEGTGRLFFEFYRLLHDARPKEGDDRPFFWLFENVVAMGVS
                     DKRDISRFLESNPVMIDAKEVSAAHRARYFWGNLPGMNRPLASTVNDKLELQECLEHGR
                     IAKFSKVRTITTRSNSIKQGKDQHFPVFMNEKEDILWCTEMERVFGFPVHYTDVSNMSR
                     LARQRLLGRSWSVPVIRHLFAPLKEYFACV"
     primer_bind     complement(8210..8227)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     8437..8456
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     CDS             8618..9328
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(8763..8781)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     9031..9050
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    9361..9949
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(9414..9434)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9832..9843)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(9952..9968)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9953..9969)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             10474..10654
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10680..10697)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10686..10910
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10956..11384
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(11043..11062)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     11253..11274
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(11393..11413)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        11398..11727
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      11578..11713
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11640..11659
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11775..11822
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11841..12215
                     /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    12345..12466
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(12382..12401)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     12436..12455
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(12515..12531)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(12515..12531)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12528..12550)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12539..12555
                     /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(12563..12593)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12608..12629
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12746..12763)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12917..13505)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12997..13016)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(13676..14536)
                     /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     14299..14318
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14537..14641)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(14716..14735)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aattgcatga agaatctgct tagggttagg cgttttgcgc tgcttcgcga tgtacgggcc
       61 agatatacgc gttgacattg attattgact agttattaat agtaatcaat tacggggtca
      121 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
      181 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
      241 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
      301 ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt
      361 aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag
      421 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacatcaat
      481 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
      541 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa caactccgcc
      601 ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagcgcgttt
      661 tgcctgtact gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
      721 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
      781 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
      841 aatctctagc agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc
      901 tctcgacgca ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg
      961 gtgagtacgc caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc
     1021 gtcagtatta agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg
     1081 ggaaagaaaa aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc
     1141 gcagttaatc ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta
     1201 caaccatccc ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc
     1261 ctctattgtg tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata
     1321 gaggaagagc aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc
     1381 tggaggagga gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa
     1441 aattgaacca ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa
     1501 aagagcagtg ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat
     1561 gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca
     1621 gcagcagaac aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt
     1681 ctggggcatc aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca
     1741 acagctcctg gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg
     1801 gaatgctagt tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga
     1861 gtgggacaga gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca
     1921 aaaccagcaa gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg
     1981 gaattggttt aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg
     2041 aggcttggta ggtttaagaa tagtttttgc tgtactttct atagtgaata gagttaggca
     2101 gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac ccgacaggcc
     2161 cgaaggaata gaagaagaag gtggagagag agacagagac agatccattc gattagtgaa
     2221 cggatcggca ctgcgtgcgc caattctgca gacaaatggc agtattcatc cacaatttta
     2281 aaagaaaagg ggggattggg gggtacagtg caggggaaag aatagtagac ataatagcaa
     2341 cagacataca aactaaagaa ttacaaaaac aaattacaaa aattcaaaat tttcgggttt
     2401 attacaggga cagcagagat ccagtttggt tagatctcga gtttaccact ccctatcagt
     2461 gatagagaaa agtgaaagtc gagtttacca ctccctatca gtgatagaga aaagtgaaag
     2521 tcgagtttac cactccctat cagtgataga gaaaagtgaa agtcgagttt accactccct
     2581 atcagtgata gagaaaagtg aaagtcgagt ttaccactcc ctatcagtga tagagaaaag
     2641 tgaaagtcga gtttaccact ccctatcagt gatagagaaa agtgaaagtc gagtttacca
     2701 ctccctatca gtgatagaga aaagtgaaag tcgagctcgg tacccgggtc gaggtaggcg
     2761 tgtacggtgg gaggcctata taagcagagc tcgtttagtg aaccgtcaga tcgcctggag
     2821 acgccatcca cgctgttttg acctccatag aagacaccgg gaccgatcca gcctccgcgg
     2881 ccccgaattc gccatacatt ctagaggatc cgccaccatg gacaagaagt actccattgg
     2941 gctcgctatc ggcacaaaca gcgtcggctg ggccgtcatt acggacgagt acaaggtgcc
     3001 gagcaaaaaa ttcaaagttc tgggcaatac cgatcgccac agcataaaga agaacctcat
     3061 tggcgccctc ctgttcgact ccggggagac ggccgaagcc acgcggctca aaagaacagc
     3121 acggcgcaga tatacccgca gaaagaatcg gatctgctac ctgcaggaga tctttagtaa
     3181 tgagatagct aaggtggatg actctttctt ccataggctg gaggagtcct ttttggtgga
     3241 ggaggataaa aagcacgagc gccacccaat ctttggcaat atcgtggacg aggtggcgta
     3301 ccatgaaaag tacccaacca tatatcatct gaggaagaag cttgtagaca gtactgataa
     3361 ggctgacttg cggttgatct atctcgcgct ggcgcatatg atcaaatttc ggggacactt
     3421 cctcatcgag ggggacctga acccagacaa cagcgatgtc gacaaactct ttatccaact
     3481 ggttcagact tacaatcagc ttttcgaaga gaacccgatc aacgcatccg gagttgacgc
     3541 caaagcaatc ctgagcgcta ggctgtccaa atcccggcgg ctcgaaaacc tcatcgcaca
     3601 gctccctggg gagaagaaga acggcctgtt tggtaatctt atcgccctgt cactcgggct
     3661 gacccccaac tttaaatcta acttcgacct ggccgaagat gccaagcttc aactgagcaa
     3721 agacacctac gatgatgatc tcgacaatct gctggcccag atcggcgacc agtacgcaga
     3781 cctttttttg gcggcaaaga acctgtcaga cgccattctg ctgagtgata ttctgcgagt
     3841 gaacacggag atcaccaaag ctccgctgag cgctagtatg atcaagcgct atgatgagca
     3901 ccaccaagac ttgactttgc tgaaggccct tgtcagacag caactgcctg agaagtacaa
     3961 ggaaattttc ttcgatcagt ctaaaaatgg ctacgccgga tacattgacg gcggagcaag
     4021 ccaggaggaa ttttacaaat ttattaagcc catcttggaa aaaatggacg gcaccgagga
     4081 gctgctggta aagcttaaca gagaagatct gttgcgcaaa cagcgcactt tcgacaatgg
     4141 aagcatcccc caccagattc acctgggcga actgcacgct atcctcaggc ggcaagagga
     4201 tttctacccc tttttgaaag ataacaggga aaagattgag aaaatcctca catttcggat
     4261 accctactat gtaggccccc tcgcccgggg aaattccaga ttcgcgtgga tgactcgcaa
     4321 atcagaagag accatcactc cctggaactt cgaggaagtc gtggataagg gggcctctgc
     4381 ccagtccttc atcgaaagga tgactaactt tgataaaaat ctgcctaacg aaaaggtgct
     4441 tcctaaacac tctctgctgt acgagtactt cacagtttat aacgagctca ccaaggtcaa
     4501 atacgtcaca gaagggatga gaaagccagc attcctgtct ggagagcaga agaaagctat
     4561 cgtggacctc ctcttcaaga cgaaccggaa agttaccgtg aaacagctca aagaagacta
     4621 tttcaaaaag attgaatgtt tcgactctgt tgaaatcagc ggagtggagg atcgcttcaa
     4681 cgcatccctg ggaacgtatc acgatctcct gaaaatcatt aaagacaagg acttcctgga
     4741 caatgaggag aacgaggaca ttcttgagga cattgtcctc acccttacgt tgtttgaaga
     4801 tagggagatg attgaagaac gcttgaaaac ttacgctcat ctcttcgacg acaaagtcat
     4861 gaaacagctc aagaggcgcc gatatacagg atgggggcgg ctgtcaagaa aactgatcaa
     4921 tgggatccga gacaagcaga gtggaaagac aatcctggat tttcttaagt ccgatggatt
     4981 tgccaacagg aacttcatgc agttgatcca tgatgactct ctcaccttta aggaggacat
     5041 ccagaaagca caagtttctg gccaggggga cagtcttcac gagcacatcg ctaatcttgc
     5101 aggtagccca gctatcaaaa agggaatact gcagaccgtt aaggtcgtgg atgaactcgt
     5161 caaagtaatg ggaaggcata agcccgagaa tatcgttatc gagatggccc gagagaacca
     5221 aactacccag aagggacaga agaacagtag ggaaaggatg aagaggattg aagagggtat
     5281 aaaagaactg gggtcccaaa tccttaagga acacccagtt gaaaacaccc agcttcagaa
     5341 tgagaagctc tacctgtact acctgcagaa cggcagggac atgtacgtgg atcaggaact
     5401 ggacatcaat cggctctccg actacgacgt ggatgctatc gtgccccagt cttttctcaa
     5461 agatgattct attgataata aagtgttgac aagatccgat aaaaatagag ggaagagcga
     5521 taacgtcccc tcagaagaag ttgtcaagaa aatgaaaaat tattggcggc agctgctgaa
     5581 cgccaaactg atcacacaac ggaagttcga taatctgact aaggctgaac gaggtggcct
     5641 gtctgagttg gataaagccg gcttcatcaa aaggcagctt gttgagacac gccagatcac
     5701 caagcacgtg gcccaaattc tcgattcacg catgaacacc aagtacgatg aaaatgacaa
     5761 actgattcga gaggtgaaag ttattactct gaagtctaag ctggtctcag atttcagaaa
     5821 ggactttcag ttttataagg tgagagagat caacaattac caccatgcgc atgatgccta
     5881 cctgaatgca gtggtaggca ctgcacttat caaaaaatat cccaagcttg aatctgaatt
     5941 tgtttacgga gactataaag tgtacgatgt taggaaaatg atcgcaaagt ctgagcagga
     6001 aataggcaag gccaccgcta agtacttctt ttacagcaat attatgaatt ttttcaagac
     6061 cgagattaca ctggccaatg gagagattcg gaagcgacca cttatcgaaa caaacggaga
     6121 aacaggagaa atcgtgtggg acaagggtag ggatttcgcg acagtccgga aggtcctgtc
     6181 catgccgcag gtgaacatcg ttaaaaagac cgaagtacag accggaggct tctccaagga
     6241 aagtatcctc ccgaaaagga acagcgacaa gctgatcgca cgcaaaaaag attgggaccc
     6301 caagaaatac ggcggattcg attctcctac agtcgcttac agtgtactgg ttgtggccaa
     6361 agtggagaaa gggaagtcta aaaaactcaa aagcgtcaag gaactgctgg gcatcgcaat
     6421 catggagcga tcaagcttcg aaaaaaaccc catcgacttt ctcgaggcga aaggatataa
     6481 agaggtcaaa aaagacctca tcattaagct tcccaagtac tctctctttg agcttgaaaa
     6541 cggccggaaa cgaatgctcg ctagtgcggg cgagctgcag aaaggtaacg agctggcact
     6601 gccctctaaa tacgttaatt tcttgtatct ggccagccac tatgaaaagc tcaaagggtc
     6661 tcccgaagat aatgagcaga agcagctgtt cgtggaacaa cacaaacact accttgatga
     6721 gatcatcgag caaataagcg aattctccaa aagagtgatc ctcgccgacg ctaacctcga
     6781 taaggtgctt tctgcttaca ataagcacag ggataagccc atcagggagc aggcagaaaa
     6841 cattatccac ttgtttactc tgaccaactt gggcgcgcct gcagccttca agtacttcga
     6901 caccaccata gacagaaagc ggtacacctc tacgaaggag gtcctggacg ccacactgat
     6961 tcatcagtca attacggggc tctatgaaac aagaatcgac ctctctcagc tcggtggaga
     7021 cagcagggct gaccccaaga agaagaggaa ggtgggagct agcgactaca aagacgatga
     7081 cgacaaggga agccgagagg actggccctc ccggctccag atgttcttcg ctaataacca
     7141 cgaccaggaa tttgaccctc caaaggttta cccacctgtc ccagctgaga agaggaagcc
     7201 catccgggtg ctgtctctct ttgatggaat cgctacaggg ctcctggtgc tgaaggactt
     7261 gggcattcag gtggaccgct acattgcctc ggaggtgtgt gaggactcca tcacggtggg
     7321 catggtgcgg caccagggga agatcatgta cgtcggggac gtccgcagcg tcacacagaa
     7381 gcatatccag gagtggggcc cattcgatct ggtgattggg ggcagtccct gcaatgacct
     7441 ctccatcgtc aaccctgctc gcaagggcct ctacgagggc actggccggc tcttctttga
     7501 gttctaccgc ctcctgcatg atgcgcggcc caaggaggga gatgatcgcc ccttcttctg
     7561 gctctttgag aatgtggtgg ccatgggcgt tagtgacaag agggacatct cgcgatttct
     7621 cgagtccaac cctgtgatga ttgatgccaa agaagtgtca gctgcacaca gggcccgcta
     7681 cttctggggt aaccttcccg gtatgaacag gccgttggca tccactgtga atgataagct
     7741 ggagctgcag gagtgtctgg agcatggcag gatagccaag ttcagcaaag tgaggaccat
     7801 tactacgagg tcaaactcca taaagcaggg caaagaccag cattttcctg tcttcatgaa
     7861 tgagaaagag gacatcttat ggtgcactga aatggaaagg gtatttggtt tcccagtcca
     7921 ctatactgac gtctccaaca tgagccgctt ggcgaggcag agactgctgg gccggtcatg
     7981 gagcgtgcca gtcatccgcc acctcttcgc tccgctgaag gagtattttg cgtgtgtgta
     8041 aaccggtact cgaccccccc ccctaacgtt actggccgaa gccgcttgga ataaggccgg
     8101 tgtgcgcttg tctatatgtt attttccacc atattgccgt cttttggcaa tgtgagggcc
     8161 cggaaacctg gccctgtctt cttgacgagc attcctaggg gtctttcccc tctcgccaaa
     8221 ggaatgcaag gtctgttgaa tgtcgtgaag gaagcagttc ctctggaagc ttcttgaaga
     8281 caaacaacgt ctgtagcgac cctttgcagg cagcggaacc ccccacctgg cgacaggtgc
     8341 ccctgcggcc aaaagccacg tgtataagat acacctgcaa aggcggcaca accccagtgc
     8401 cacgttgtga gttggatagt tgtggaaaga gtcaaatggc tctcctcaag cgtattcaac
     8461 aaggggctga aggatgccca gaaggtaccc cattgtatgg gatctgatct ggggcctcgg
     8521 tgcacatgct ttacatgtgt ttagtcgagg ttaaaaaaac gtctaggccc cccgaaccac
     8581 ggggacgtgg ttttcctttg aaaaacacga taataccatg gtgagcaagg gcgaggagga
     8641 taacatggcc atcatcaagg agttcatgcg cttcaaggtg cacatggagg gctccgtgaa
     8701 cggccacgag ttcgagatcg agggcgaggg cgagggccgc ccctacgagg gcacccagac
     8761 cgccaagctg aaggtgacca agggtggccc cctgcccttc gcctgggaca tcctgtcccc
     8821 tcagttcatg tacggctcca aggcctacgt gaagcacccc gccgacatcc ccgactactt
     8881 gaagctgtcc ttccccgagg gcttcaagtg ggagcgcgtg atgaacttcg aggacggcgg
     8941 cgtggtgacc gtgacccagg actcctccct gcaggacggc gagttcatct acaaggtgaa
     9001 gctgcgcggc accaacttcc cctccgacgg ccccgtaatg cagaagaaga ccatgggctg
     9061 ggaggcctcc tccgagcgga tgtaccccga ggacggcgcc ctgaagggcg agatcaagca
     9121 gaggctgaag ctgaaggacg gcggccacta cgacgctgag gtcaagacca cctacaaggc
     9181 caagaagccc gtgcagctgc ccggcgccta caacgtcaac atcaagttgg acatcacctc
     9241 ccacaacgag gactacacca tcgtggaaca gtacgaacgc gccgagggcc gccactccac
     9301 cggcggcatg gacgagctgt acaagtagac gcgcccgaat tcgatatcaa gcttatcgat
     9361 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     9421 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     9481 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     9541 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     9601 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     9661 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     9721 ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc
     9781 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     9841 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     9901 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt
     9961 cgacctcgag acctagaaaa acatggagca atcacaagta gcaatacagc agctaccaat
    10021 gctgattgtg cctggctaga agcacaagag gaggaggagg tgggttttcc agtcacacct
    10081 caggtacctt taagaccaat gacttacaag gcagctgtag atcttagcca ctttttaaaa
    10141 gaaaaggggg gactggaagg gctaattcac tcccaacgaa gacaagatat ccttgatctg
    10201 tggatctacc acacacaagg ctacttccct gattggcaga actacacacc agggccaggg
    10261 atcagatatc cactgacctt tggatggtgc tacaagctag taccagttga gcaagagaag
    10321 gtagaagaag ccaatgaagg agagaacacc cgcttgttac accctgtgag cctgcatggg
    10381 atggatgacc cggagagaga agtattagag tggaggtttg acagccgcct agcatttcat
    10441 cacatggccc gagagctgca tccggactgt actgggtctc tctggttaga ccagatctga
    10501 gcctgggagc tctctggcta actagggaac ccactgctta agcctcaata aagcttgcct
    10561 tgagtgcttc aagtagtgtg tgcccgtctg ttgtgtgact ctggtaacta gagatccctc
    10621 agaccctttt agtcagtgtg gaaaatctct agcagggccc gtttaaaccc gctgatcagc
    10681 ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg tgccttcctt
    10741 gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa ttgcatcgca
    10801 ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca gcaaggggga
    10861 ggattgggaa gacaatagca ggcatgctgg ggatgcggtg ggctctatgg cttctgaggc
    10921 ggaaagaacc agctggggct ctagggggta tccccacgcg ccctgtagcg gcgcattaag
    10981 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
    11041 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
    11101 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
    11161 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
    11221 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
    11281 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
    11341 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattaattct gtggaatgtg
    11401 tgtcagttag ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg
    11461 catctcaatt agtcagcaac caggtgtgga aagtccccag gctccccagc aggcagaagt
    11521 atgcaaagca tgcatctcaa ttagtcagca accatagtcc cgcccctaac tccgcccatc
    11581 ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact aatttttttt
    11641 atttatgcag aggccgaggc cgcctctgcc tctgagctat tccagaagta gtgaggaggc
    11701 ttttttggag gcctaggctt ttgcaaaaag ctcccgggag cttgtatatc cattttcgga
    11761 tctgatcagc acgtgttgac aattaatcat cggcatagta tatcggcata gtataatacg
    11821 acaaggtgag gaactaaacc atggccaagt tgaccagtgc cgttccggtg ctcaccgcgc
    11881 gcgacgtcgc cggagcggtc gagttctgga ccgaccggct cgggttctcc cgggacttcg
    11941 tggaggacga cttcgccggt gtggtccggg acgacgtgac cctgttcatc agcgcggtcc
    12001 aggaccaggt ggtgccggac aacaccctgg cctgggtgtg ggtgcgcggc ctggacgagc
    12061 tgtacgccga gtggtcggag gtcgtgtcca cgaacttccg ggacgcctcc gggccggcca
    12121 tgaccgagat cggcgagcag ccgtgggggc gggagttcgc cctgcgcgac ccggccggca
    12181 actgcgtgca cttcgtggcc gaggagcagg actgacacgt gctacgagat ttcgattcca
    12241 ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt ccgggacgcc ggctggatga
    12301 tcctccagcg cggggatctc atgctggagt tcttcgccca ccccaacttg tttattgcag
    12361 cttataatgg ttacaaataa agcaatagca tcacaaattt cacaaataaa gcattttttt
    12421 cactgcattc tagttgtggt ttgtccaaac tcatcaatgt atcttatcat gtctgtatac
    12481 cgtcgacctc tagctagagc ttggcgtaat catggtcata gctgtttcct gtgtgaaatt
    12541 gttatccgct cacaattcca cacaacatac gagccggaag cataaagtgt aaagcctggg
    12601 gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg ctcactgccc gctttccagt
    12661 cgggaaacct gtcgtgccag ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
    12721 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
    12781 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
    12841 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
    12901 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
    12961 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
    13021 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
    13081 ttctcccttc gggaagcgtg gcgctttctc atagctcacg ctgtaggtat ctcagttcgg
    13141 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
    13201 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
    13261 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
    13321 tcttgaagtg gtggcctaac tacggctaca ctagaagaac agtatttggt atctgcgctc
    13381 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
    13441 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
    13501 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
    13561 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
    13621 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
    13681 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
    13741 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
    13801 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
    13861 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
    13921 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
    13981 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
    14041 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
    14101 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
    14161 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
    14221 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
    14281 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
    14341 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
    14401 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
    14461 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
    14521 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
    14581 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
    14641 gcacatttcc ccgaaaagtg ccacctgacg tcgacggatc gggagatctc ccgatcccct
    14701 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atctgctccc
    14761 tgcttgtgtg ttggaggtcg ctgagtagtg cgcgagcaaa atttaagcta caacaaggca
    14821 aggcttgacc gac
//
