LOCUS       sequence-197882-        9265 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  transient expression of dCas9-VP64 fusion protein.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9265)
  AUTHORS   Stephan Beck
  TITLE     Beck lab Cas9 plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9265)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     Sequence Label: p-dCas9-VP64-Hygro
FEATURES             Location/Qualifiers
     source          1..9265
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     polyA_signal    86..207
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(97..116)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     151..170
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(238..1263)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     CDS             complement(238..1263)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     promoter        complement(1281..1780)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(1329..1348)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     1722..1745
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     enhancer        1785..2164
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2165..2368
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2319..2339
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2410..2429
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2410..2428
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      2436..2445
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2442..5843,5844..5846,5847..6443,6444,6445..6447,
                     6448..6449,6450..6452,6453..6545)
                     /codon_start=1
                     /product="Streptococcus pyogenes Cas9 endonuclease with the
                     D1135V/R1335Q/T1337R mutations"
                     /label=Cas9 VQR
                     /note="has an altered recognition specificity for the 
                     protospacer adjacent motif (PAM) (Kleinstiver et al., 
                     2015)"
                     /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             join(2442..2468,2469..2471,2472..4958,4959..4961,
                     4962..6545)
                     /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             6552..6572
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             6579..6644
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             6675..6824
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             6833..6850
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6873..6890)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6879..7103
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(7174..7190)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7174..7190)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7187..7209)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7198..7214
                     /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(7222..7228,7229..7246,7247..7252))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7267..7288
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7405..7422)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7576..8164)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7656..7675)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(8335..9126,9127..9195))
                     /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     8958..8977
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(9196..9265,1..35))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
       61 gacgtcgacg ggtatacaga catgataaga tacattgatg agtttggaca aaccacaact
      121 agaatgcagt gaaaaaaatg ctttatttgt gaaatttgtg atgctattgc tttatttgta
      181 accattataa gctgcaataa acaagttggg gtgggcgaag aactctcggc atctactcta
      241 ttcctttgcc ctcggacgag tgctggggcg tcggtttcca ctatcggcga gtacttctac
      301 acagccatcg gtccagacgg ccgcgcttct gcgggcgatt tgtgtacgcc cgacagtccc
      361 ggctccggat cggacgattg cgtcgcatcg accctgcgcc caagctgcat catcgaaatt
      421 gccgtcaacc aagctctgat agagttggtc aagaccaatg cggagcatat acgcccggag
      481 ccgcggcgat cctgcaagct ccggatgcct ccgctcgaag tagcgcgtct gctgctccat
      541 acaagccaac cacggcctcc agaagaagat gttggcgacc tcgtattggg aatccccgaa
      601 catcgcctcg ctccagtcaa tgaccgctgt tatgcggcca ttgtccgtca ggacattgtt
      661 ggagccgaaa tccgcgtgca cgaggtgccg gacttcgggg cagtcctcgg cccaaagcat
      721 cagctcatcg agagcctgcg cgacggacgc actgacggtg tcgtccatca cagtttgcca
      781 gtgatacaca tggggatcag caatcgcgca tatgaaatca cgccatgtag tgtattgacc
      841 gattccttgc ggtccgaatg ggccgaaccc gctcgtctgg ctaagatcgg ccgcagcgat
      901 cgcatccatg gcctccgcga ccggctgcag aacagcgggc agttcggttt caggcaggtc
      961 ttgcaacgtg acaccctgtg cacggcggga gatgcaatag gtcaggctct cgctgaattc
     1021 cccaatgtca agcacttccg gaatcgggag cgcggccgat gcaaagtgcc gataaacata
     1081 acgatctttg tagaaaccat cggcgcagct atttacccgc aggacatatc cacgccctcc
     1141 tacatcgaag ctgaaagcac gagattcttc gccctccgag agctgcatca ggtcggagac
     1201 gctgtcgaac ttttcgatca gaaacttctc gacagacgtc gcggtgagtt caggcttttt
     1261 catggtggcg ggatgcaggt cgaaaggccc ggagatgagg aagaggagaa cagcgcggca
     1321 gacgtgcgct tttgaagcgt gcagaatgcc gggcctccgg aggaccttcg ggcgcccgcc
     1381 ccgcccctga gcccgcccct gagcccgccc ccggacccac cccttcccag cctctgagcc
     1441 cagaaagcga aggagcaaag ctgctattgg ccgctgcccc aaaggcctac ccgcttccat
     1501 tgctcagcgg tgctgtccat ctgcacgaga ctagtgagac gtgctacttc catttgtcac
     1561 gtcctgcacg acgcgagctg cggggcgggg gggaacttcc tgactagggg aggagtagaa
     1621 ggtggcgcga aggggccacc aaagaacgga gccggttggc gcctaccggt ggatgtggaa
     1681 tgtgtgcgag gccagaggcc acttgtgtag cgccaagtgc ccagcggggc tgctaaagcg
     1741 catgctccag actgccttgg gaaaagcgcc tcccctaccc ggttgacatt gattattgac
     1801 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
     1861 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
     1921 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
     1981 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
     2041 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
     2101 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
     2161 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
     2221 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
     2281 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
     2341 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagagatc
     2401 cgcggccgct aatacgactc actataggga gagccgccac catggataaa aagtattcta
     2461 ttggtttagc catcggcact aattccgttg gatgggctgt cataaccgat gaatacaaag
     2521 taccttcaaa gaaatttaag gtgttgggga acacagaccg tcattcgatt aaaaagaatc
     2581 ttatcggtgc cctcctattc gatagtggcg aaacggcaga ggcgactcgc ctgaaacgaa
     2641 ccgctcggag aaggtataca cgtcgcaaga accgaatatg ttacttacaa gaaattttta
     2701 gcaatgagat ggccaaagtt gacgattctt tctttcaccg tttggaagag tccttccttg
     2761 tcgaagagga caagaaacat gaacggcacc ccatctttgg aaacatagta gatgaggtgg
     2821 catatcatga aaagtaccca acgatttatc acctcagaaa aaagctagtt gactcaactg
     2881 ataaagcgga cctgaggtta atctacttgg ctcttgccca tatgataaag ttccgtgggc
     2941 actttctcat tgagggtgat ctaaatccgg acaactcgga tgtcgacaaa ctgttcatcc
     3001 agttagtaca aacctataat cagttgtttg aagagaaccc tataaatgca agtggcgtgg
     3061 atgcgaaggc tattcttagc gcccgcctct ctaaatcccg acggctagaa aacctgatcg
     3121 cacaattacc cggagagaag aaaaatgggt tgttcggtaa ccttatagcg ctctcactag
     3181 gcctgacacc aaattttaag tcgaacttcg acttagctga agatgccaaa ttgcagctta
     3241 gtaaggacac gtacgatgac gatctcgaca atctactggc acaaattgga gatcagtatg
     3301 cggacttatt tttggctgcc aaaaacctta gcgatgcaat cctcctatct gacatactga
     3361 gagttaatac tgagattacc aaggcgccgt tatccgcttc aatgatcaaa aggtacgatg
     3421 aacatcacca agacttgaca cttctcaagg ccctagtccg tcagcaactg cctgagaaat
     3481 ataaggaaat attctttgat cagtcgaaaa acgggtacgc aggttatatt gacggcggag
     3541 cgagtcaaga ggaattctac aagtttatca aacccatatt agagaagatg gatgggacgg
     3601 aagagttgct tgtaaaactc aatcgcgaag atctactgcg aaagcagcgg actttcgaca
     3661 acggtagcat tccacatcaa atccacttag gcgaattgca tgctatactt agaaggcagg
     3721 aggattttta tccgttcctc aaagacaatc gtgaaaagat tgagaaaatc ctaacctttc
     3781 gcatacctta ctatgtggga cccctggccc gagggaactc tcggttcgca tggatgacaa
     3841 gaaagtccga agaaacgatt actccatgga attttgagga agttgtcgat aaaggtgcgt
     3901 cagctcaatc gttcatcgag aggatgacca actttgacaa gaatttaccg aacgaaaaag
     3961 tattgcctaa gcacagttta ctttacgagt atttcacagt gtacaatgaa ctcacgaaag
     4021 ttaagtatgt cactgagggc atgcgtaaac ccgcctttct aagcggagaa cagaagaaag
     4081 caatagtaga tctgttattc aagaccaacc gcaaagtgac agttaagcaa ttgaaagagg
     4141 actactttaa gaaaattgaa tgcttcgatt ctgtcgagat ctccggggta gaagatcgat
     4201 ttaatgcgtc acttggtacg tatcatgacc tcctaaagat aattaaagat aaggacttcc
     4261 tggataacga agagaatgaa gatatcttag aagatatagt gttgactctt accctctttg
     4321 aagatcggga aatgattgag gaaagactaa aaacatacgc tcacctgttc gacgataagg
     4381 ttatgaaaca gttaaagagg cgtcgctata cgggctgggg acgattgtcg cggaaactta
     4441 tcaacgggat aagagacaag caaagtggta aaactattct cgattttcta aagagcgacg
     4501 gcttcgccaa taggaacttt atgcagctga tccatgatga ctctttaacc ttcaaagagg
     4561 atatacaaaa ggcacaggtt tccggacaag gggactcatt gcacgaacat attgcgaatc
     4621 ttgctggttc gccagccatc aaaaagggca tactccagac agtcaaagta gtggatgagc
     4681 tagttaaggt catgggacgt cacaaaccgg aaaacattgt aatcgagatg gcacgcgaaa
     4741 atcaaacgac tcagaagggg caaaaaaaca gtcgagagcg gatgaagaga atagaagagg
     4801 gtattaaaga actgggcagc cagatcttaa aggagcatcc tgtggaaaat acccaattgc
     4861 agaacgagaa actttacctc tattacctac aaaatggaag ggacatgtat gttgatcagg
     4921 aactggacat aaaccgttta tctgattacg acgtcgatgc cattgtaccc caatcctttt
     4981 tgaaggacga ttcaatcgac aataaagtgc ttacacgctc ggataagaac cgagggaaaa
     5041 gtgacaatgt tccaagcgag gaagtcgtaa agaaaatgaa gaactattgg cggcagctcc
     5101 taaatgcgaa actgataacg caaagaaagt tcgataactt aactaaagct gagaggggtg
     5161 gcttgtctga acttgacaag gccggattta ttaaacgtca gctcgtggaa acccgccaaa
     5221 tcacaaagca tgttgcacag atactagatt cccgaatgaa tacgaaatac gacgagaacg
     5281 ataagctgat tcgggaagtc aaagtaatca ctttaaagtc aaaattggtg tcggacttca
     5341 gaaaggattt tcaattctat aaagttaggg agataaataa ctaccaccat gcgcacgacg
     5401 cttatcttaa tgccgtcgta gggaccgcac tcattaagaa atacccgaag ctagaaagtg
     5461 agtttgtgta tggtgattac aaagtttatg acgtccgtaa gatgatcgcg aaaagcgaac
     5521 aggagatagg caaggctaca gccaaatact tcttttattc taacattatg aatttcttta
     5581 agacggaaat cactctggca aacggagaga tacgcaaacg acctttaatt gaaaccaatg
     5641 gggagacagg tgaaatcgta tgggataagg gccgggactt cgcgacggtg agaaaagttt
     5701 tgtccatgcc ccaagtcaac atagtaaaga aaactgaggt gcagaccgga gggttttcaa
     5761 aggaatcgat tcttccaaaa aggaatagtg ataagctcat cgctcgtaaa aaggactggg
     5821 acccgaaaaa gtacggtggc ttcgatagcc ctacagttgc ctattctgtc ctagtagtgg
     5881 caaaagttga gaagggaaaa tccaagaaac tgaagtcagt caaagaatta ttggggataa
     5941 cgattatgga gcgctcgtct tttgaaaaga accccatcga cttccttgag gcgaaaggtt
     6001 acaaggaagt aaaaaaggat ctcataatta aactaccaaa gtatagtctg tttgagttag
     6061 aaaatggccg aaaacggatg ttggctagcg ccggagagct tcaaaagggg aacgaactcg
     6121 cactaccgtc taaatacgtg aatttcctgt atttagcgtc ccattacgag aagttgaaag
     6181 gttcacctga agataacgaa cagaagcaac tttttgttga gcagcacaaa cattatctcg
     6241 acgaaatcat agagcaaatt tcggaattca gtaagagagt catcctagct gatgccaatc
     6301 tggacaaagt attaagcgca tacaacaagc acagggataa acccatacgt gagcaggcgg
     6361 aaaatattat ccatttgttt actcttacca acctcggcgc tccagccgca ttcaagtatt
     6421 ttgacacaac gatagatcgc aaacgataca cttctaccaa ggaggtgcta gacgcgacac
     6481 tgattcacca atccatcacg ggattatatg aaactcggat agatttgtca cagcttgggg
     6541 gtgacggatc ccccaagaag aagaggaaag tctcgagcga ctacaaagac catgacggtg
     6601 attataaaga tcatgacatc gattacaagg atgacgatga caaggctgca ggaggcggtg
     6661 gaagcgggcg cgccgacgcg ctggacgatt tcgatctcga catgctgggt tctgatgccc
     6721 tcgatgactt tgacctggat atgttgggaa gcgacgcatt ggatgacttt gatctggaca
     6781 tgctcggctc cgatgctctg gacgatttcg atctcgatat gttataaccg gtcatcatca
     6841 ccatcaccat tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     6901 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     6961 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     7021 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     7081 tggggatgcg gtgggctcta tggcttctga ggcggaaaga accagctggg gctcgatacc
     7141 gtcgacctct agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg
     7201 ttatccgctc acaattccac acaacatacg agccggaagc ataaagtgta aagcctaggg
     7261 tgcctaatga gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc
     7321 gggaaacctg tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt
     7381 gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct
     7441 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     7501 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     7561 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     7621 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     7681 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     7741 tctcccttcg ggaagcgtgg cgctttctca atgctcacgc tgtaggtatc tcagttcggt
     7801 gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg
     7861 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     7921 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     7981 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     8041 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     8101 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     8161 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     8221 ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta
     8281 aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca
     8341 atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc
     8401 ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc
     8461 tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa taaaccagcc
     8521 agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat
     8581 taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt
     8641 tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc
     8701 cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag
     8761 ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt
     8821 tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac
     8881 tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg
     8941 cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat
     9001 tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc
     9061 gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc
     9121 tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa
     9181 atgttgaata ctcatactct tcctttttca atattattga agcatttatc agggttattg
     9241 tctcatgagc ggatacatat ttgaa
//
