LOCUS       sequence-85582-e       13567 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses taCas9 in Mammalian cells for transactivating endogenous 
            and synthetic promoters. The backbone is a lentiviral vector..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13567)
  AUTHORS   Nissim L, Perli SD, Fridkin A, Perez-Pinera P, Lu TK
  TITLE     Multiplexed and Programmable Regulation of Gene Networks with an 
            Integrated RNA and CRISPR/Cas Toolkit in Human Cells.
  JOURNAL   Mol Cell. 2014 May 14. pii: S1097-2765(14)00355-4. doi: 
            10.1016/j.molcel.2014.04.022.
  PUBMED    24837679
REFERENCE   2  (bases 1 to 13567)
  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: "Mol Cell. 
            2014 May 14. pii: S1097-2765(14)00355-4. doi: 
            10.1016/j.molcel.2014.04.022."
COMMENT     Sequence Label: CMVp-dCas9-3xNLS-VP64 (Construct 1)
FEATURES             Location/Qualifiers
     source          1..13567
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(47..66)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        238..617
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        619..817
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     772..792
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             835..1015
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1062..1187
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1680..1913
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2098..2142
                     /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    2440..2557
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2611..3821
                     /label=UbC promoter
                     /note="human ubiquitin C promoter"
     primer_bind     3658..3677
                     /label=hUBCpro-F
                     /note="Human Ubiquitin C (UbC) promoter, forward primer"
     regulatory      3824..3833
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3830..3856,3857..3859,3860..6343,6344..6349,6350..6415,
                     6416..6418,6419..7933)
                     /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
                     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(3830..3856,3857..3859,3860..6346,6347..6349,
                     6350..7933)
                     /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             3830..7933
                     /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
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             7970..7990
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7997..8017
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8024..8044
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8084..8233
                     /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"
     misc_feature    8259..8847
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8312..8332)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8730..8741)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(8850..8866)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(8851..8867)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     LTR             9372..9552
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(9578..9595)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    9584..9808
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      9854..10282
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9941..9960)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10151..10172
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10291..10311)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10296..10625
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      10476..10611
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     10538..10557
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        10673..10720
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             10739..11113
                     /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    11243..11364
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11280..11299)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11334..11353
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11413..11429)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11413..11429)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11426..11448)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11437..11453
                     /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(11461..11467,11468..11485,11486..11491))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11506..11527
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11644..11661)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11815..12403)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11895..11914)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12574..13365,13366..13434))
                     /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     13197..13216
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(13435..13539)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacggat cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg
       61 atgccgcata gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt
      121 gcgcgagcaa aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc
      181 tgcttagggt taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac
      241 attgattatt gactagttat taatagtaat caattacggg gtcattagtt catagcccat
      301 atatggagtt ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg
      361 acccccgccc attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt
      421 tccattgacg tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag
      481 tgtatcatat gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc
      541 attatgccca gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag
      601 tcatcgctat taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt
      661 ttgactcacg gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc
      721 accaaaatca acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg
      781 gcggtaggcg tgtacggtgg gaggtctata taagcagcgc gttttgcctg tactgggtct
      841 ctctggttag accagatctg agcctgggag ctctctggct aactagggaa cccactgctt
      901 aagcctcaat aaagcttgcc ttgagtgctt caagtagtgt gtgcccgtct gttgtgtgac
      961 tctggtaact agagatccct cagacccttt tagtcagtgt ggaaaatctc tagcagtggc
     1021 gcccgaacag ggacttgaaa gcgaaaggga aaccagagga gctctctcga cgcaggactc
     1081 ggcttgctga agcgcgcacg gcaagaggcg aggggcggcg actggtgagt acgccaaaaa
     1141 ttttgactag cggaggctag aaggagagag atgggtgcga gagcgtcagt attaagcggg
     1201 ggagaattag atcgcgatgg gaaaaaattc ggttaaggcc agggggaaag aaaaaatata
     1261 aattaaaaca tatagtatgg gcaagcaggg agctagaacg attcgcagtt aatcctggcc
     1321 tgttagaaac atcagaaggc tgtagacaaa tactgggaca gctacaacca tcccttcaga
     1381 caggatcaga agaacttaga tcattatata atacagtagc aaccctctat tgtgtgcatc
     1441 aaaggataga gataaaagac accaaggaag ctttagacaa gatagaggaa gagcaaaaca
     1501 aaagtaagac caccgcacag caagcggccg ctgatcttca gacctggagg aggagatatg
     1561 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1621 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     1681 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     1741 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     1801 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     1861 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     1921 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     1981 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2041 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2101 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2161 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2221 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2281 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2341 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc ggcactgcgt
     2401 gcgccaattc tgcagacaaa tggcagtatt catccacaat tttaaaagaa aaggggggat
     2461 tggggggtac agtgcagggg aaagaatagt agacataata gcaacagaca tacaaactaa
     2521 agaattacaa aaacaaatta caaaaattca aaattttcgg gtttattaca gggacagcag
     2581 agatccagtt tggttaatta agggtgcagc ggcctccgcg ccgggttttg gcgcctcccg
     2641 cgggcgcccc cctcctcacg gcgagcgctg ccacgtcaga cgaagggcgc aggagcgttc
     2701 ctgatccttc cgcccggacg ctcaggacag cggcccgctg ctcataagac tcggccttag
     2761 aaccccagta tcagcagaag gacattttag gacgggactt gggtgactct agggcactgg
     2821 ttttctttcc agagagcgga acaggcgagg aaaagtagtc ccttctcggc gattctgcgg
     2881 agggatctcc gtggggcggt gaacgccgat gattatataa ggacgcgccg ggtgtggcac
     2941 agctagttcc gtcgcagccg ggatttgggt cgcggttctt gtttgtggat cgctgtgatc
     3001 gtcacttggt gagttgcggg ctgctgggct ggccggggct ttcgtggccg ccgggccgct
     3061 cggtgggacg gaagcgtgtg gagagaccgc caagggctgt agtctgggtc cgcgagcaag
     3121 gttgccctga actgggggtt ggggggagcg cacaaaatgg cggctgttcc cgagtcttga
     3181 atggaagacg cttgtaaggc gggctgtgag gtcgttgaaa caaggtgggg ggcatggtgg
     3241 gcggcaagaa cccaaggtct tgaggccttc gctaatgcgg gaaagctctt attcgggtga
     3301 gatgggctgg ggcaccatct ggggaccctg acgtgaagtt tgtcactgac tggagaactc
     3361 gggtttgtcg tctggttgcg ggggcggcag ttatgcggtg ccgttgggca gtgcacccgt
     3421 acctttggga gcgcgcgcct cgtcgtgtcg tgacgtcacc cgttctgttg gcttataatg
     3481 cagggtgggg ccacctgccg gtaggtgtgc ggtaggcttt tctccgtcgc aggacgcagg
     3541 gttcgggcct agggtaggct ctcctgaatc gacaggcgcc ggacctctgg tgaggggagg
     3601 gataagtgag gcgtcagttt ctttggtcgg ttttatgtac ctatcttctt aagtagctga
     3661 agctccggtt ttgaactatg cgctcggggt tggcgagtgt gttttgtgaa gttttttagg
     3721 caccttttga aatgtaatca tttgggtcaa tatgtaattt tcagtgttag actagtaaat
     3781 tgtccgctaa attctggccg tttttggctt ttttgttaga caggccacca tggacaagaa
     3841 gtactccatt gggctcgcca tcggcacaaa cagcgtcggc tgggccgtca ttacggacga
     3901 gtacaaggtg ccgagcaaaa aattcaaagt tctgggcaat accgatcgcc acagcataaa
     3961 gaagaacctc attggcgccc tcctgttcga ctccggggag acggccgaag ccacgcggct
     4021 caaaagaaca gcacggcgca gatatacccg cagaaagaat cggatctgct acctgcagga
     4081 gatctttagt aatgagatgg ctaaggtgga tgactctttc ttccataggc tggaggagtc
     4141 ctttttggtg gaggaggata aaaagcacga gcgccaccca atctttggca atatcgtgga
     4201 cgaggtggcg taccatgaaa agtacccaac catatatcat ctgaggaaga agcttgtaga
     4261 cagtactgat aaggctgact tgcggttgat ctatctcgcg ctggcgcata tgatcaaatt
     4321 tcggggacac ttcctcatcg agggggacct gaacccagac aacagcgatg tcgacaaact
     4381 ctttatccaa ctggttcaga cttacaatca gcttttcgaa gagaacccga tcaacgcatc
     4441 cggagttgac gccaaagcaa tcctgagcgc taggctgtcc aaatcccggc ggctcgaaaa
     4501 cctcatcgca cagctccctg gggagaagaa gaacggcctg tttggtaatc ttatcgccct
     4561 gtcactcggg ctgaccccca actttaaatc taacttcgac ctggccgaag atgccaagct
     4621 tcaactgagc aaagacacct acgatgatga tctcgacaat ctgctggccc agatcggcga
     4681 ccagtacgca gacctttttt tggcggcaaa gaacctgtca gacgccattc tgctgagtga
     4741 tattctgcga gtgaacacgg agatcaccaa agctccgctg agcgctagta tgatcaagcg
     4801 ctatgatgag caccaccaag acttgacttt gctgaaggcc cttgtcagac agcaactgcc
     4861 tgagaagtac aaggaaattt tcttcgatca gtctaaaaat ggctacgccg gatacattga
     4921 cggcggagca agccaggagg aattttacaa atttattaag cccatcttgg aaaaaatgga
     4981 cggcaccgag gagctgctgg taaagcttaa cagagaagat ctgttgcgca aacagcgcac
     5041 tttcgacaat ggaagcatcc cccaccagat tcacctgggc gaactgcacg ctatcctcag
     5101 gcggcaagag gatttctacc cctttttgaa agataacagg gaaaagattg agaaaatcct
     5161 cacatttcgg ataccctact atgtaggccc cctcgcccgg ggaaattcca gattcgcgtg
     5221 gatgactcgc aaatcagaag agaccatcac tccctggaac ttcgaggaag tcgtggataa
     5281 gggggcctct gcccagtcct tcatcgaaag gatgactaac tttgataaaa atctgcctaa
     5341 cgaaaaggtg cttcctaaac actctctgct gtacgagtac ttcacagttt ataacgagct
     5401 caccaaggtc aaatacgtca cagaagggat gagaaagcca gcattcctgt ctggagagca
     5461 gaagaaagct atcgtggacc tcctcttcaa gacgaaccgg aaagttaccg tgaaacagct
     5521 caaagaagac tatttcaaaa agattgaatg tttcgactct gttgaaatca gcggagtgga
     5581 ggatcgcttc aacgcatccc tgggaacgta tcacgatctc ctgaaaatca ttaaagacaa
     5641 ggacttcctg gacaatgagg agaacgagga cattcttgag gacattgtcc tcacccttac
     5701 gttgtttgaa gatagggaga tgattgaaga acgcttgaaa acttacgctc atctcttcga
     5761 cgacaaagtc atgaaacagc tcaagaggcg ccgatataca ggatgggggc ggctgtcaag
     5821 aaaactgatc aatgggatcc gagacaagca gagtggaaag acaatcctgg attttcttaa
     5881 gtccgatgga tttgccaacc ggaacttcat gcagttgatc catgatgact ctctcacctt
     5941 taaggaggac atccagaaag cacaagtttc tggccagggg gacagtcttc acgagcacat
     6001 cgctaatctt gcaggtagcc cagctatcaa aaagggaata ctgcagaccg ttaaggtcgt
     6061 ggatgaactc gtcaaagtaa tgggaaggca taagcccgag aatatcgtta tcgagatggc
     6121 ccgagagaac caaactaccc agaagggaca gaagaacagt agggaaagga tgaagaggat
     6181 tgaagagggt ataaaagaac tggggtccca aatccttaag gaacacccag ttgaaaacac
     6241 ccagcttcag aatgagaagc tctacctgta ctacctgcag aacggcaggg acatgtacgt
     6301 ggatcaggaa ctggacatca atcggctctc cgactacgac gtggatgcca tcgtgcccca
     6361 gtcttttctc aaagatgatt ctattgataa taaagtgttg acaagatccg ataaaaatag
     6421 agggaagagt gataacgtcc cctcagaaga agttgtcaag aaaatgaaaa attattggcg
     6481 gcagctgctg aacgccaaac tgatcacaca acggaagttc gataatctga ctaaggctga
     6541 acgaggtggc ctgtctgagt tggataaagc cggcttcatc aaaaggcagc ttgttgagac
     6601 acgccagatc accaagcacg tggcccaaat tctcgattca cgcatgaaca ccaagtacga
     6661 tgaaaatgac aaactgattc gagaggtgaa agttattact ctgaagtcta agctggtctc
     6721 agatttcaga aaggactttc agttttataa ggtgagagag atcaacaatt accaccatgc
     6781 gcatgatgcc tacctgaatg cagtggtagg cactgcactt atcaaaaaat atcccaagct
     6841 tgaatctgaa tttgtttacg gagactataa agtgtacgat gttaggaaaa tgatcgcaaa
     6901 gtctgagcag gaaataggca aggccaccgc taagtacttc ttttacagca atattatgaa
     6961 ttttttcaag accgagatta cactggccaa tggagagatt cggaagcgac cacttatcga
     7021 aacaaacgga gaaacaggag aaatcgtgtg ggacaagggt agggatttcg cgacagtccg
     7081 gaaggtcctg tccatgccgc aggtgaacat cgttaaaaag accgaagtac agaccggagg
     7141 cttctccaag gaaagtatcc tcccgaaaag gaacagcgac aagctgatcg cacgcaaaaa
     7201 agattgggac cccaagaaat acggcggatt cgattctcct acagtcgctt acagtgtact
     7261 ggttgtggcc aaagtggaga aagggaagtc taaaaaactc aaaagcgtca aggaactgct
     7321 gggcatcaca atcatggagc gatcaagctt cgaaaaaaac cccatcgact ttctcgaggc
     7381 gaaaggatat aaagaggtca aaaaagacct catcattaag cttcccaagt actctctctt
     7441 tgagcttgaa aacggccgga aacgaatgct cgctagtgcg ggcgagctgc agaaaggtaa
     7501 cgagctggca ctgccctcta aatacgttaa tttcttgtat ctggccagcc actatgaaaa
     7561 gctcaaaggg tctcccgaag ataatgagca gaagcagctg ttcgtggaac aacacaaaca
     7621 ctaccttgat gagatcatcg agcaaataag cgaattctcc aaaagagtga tcctcgccga
     7681 cgctaacctc gataaggtgc tttctgctta caataagcac agggataagc ccatcaggga
     7741 gcaggcagaa aacattatcc acttgtttac tctgaccaac ttgggcgcgc ctgcagcctt
     7801 caagtacttc gacaccacca tagacagaaa gcggtacacc tctacaaagg aggtcctgga
     7861 cgccacactg attcatcagt caattacggg gctctatgaa acaagaatcg acctctctca
     7921 gctcggtgga gacagcaggg ctgacgggcc ctcactgggt tcagggtcac ccaagaagaa
     7981 acgcaaagtc gaggatccaa agaagaaaag gaaggttgaa gaccccaaga aaaagaggaa
     8041 ggtggatggg atcggctcag gcagcaacgg cggtggaggt tcagacgctt tggacgattt
     8101 cgatctcgat atgctcggtt ctgacgccct ggatgatttc gatctggata tgctcggcag
     8161 cgacgctctc gacgatttcg acctcgacat gctcgggtca gatgccttgg atgattttga
     8221 cctggatatg ctctcatgac gatatcaagc ttatcgataa tcaacctctg gattacaaaa
     8281 tttgtgaaag attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg
     8341 ctgctttaat gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct
     8401 tgtataaatc ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg
     8461 gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct
     8521 gtcagctcct ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg
     8581 ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg
     8641 tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc ctgtgttgcc acctggattc
     8701 tgcgcgggac gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc
     8761 gcggcctgct gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc
     8821 ggatctccct ttgggccgcc tccccgcatc gataccgtcg acctcgagac ctagaaaaac
     8881 atggagcaat cacaagtagc aatacagcag ctaccaatgc tgattgtgcc tggctagaag
     8941 cacaagagga ggaggaggtg ggttttccag tcacacctca ggtaccttta agaccaatga
     9001 cttacaaggc agctgtagat cttagccact ttttaaaaga aaagggggga ctggaagggc
     9061 taattcactc ccaacgaaga caagatatcc ttgatctgtg gatctaccac acacaaggct
     9121 acttccctga ttggcagaac tacacaccag ggccagggat cagatatcca ctgacctttg
     9181 gatggtgcta caagctagta ccagttgagc aagagaaggt agaagaagcc aatgaaggag
     9241 agaacacccg cttgttacac cctgtgagcc tgcatgggat ggatgacccg gagagagaag
     9301 tattagagtg gaggtttgac agccgcctag catttcatca catggcccga gagctgcatc
     9361 cggactgtac tgggtctctc tggttagacc agatctgagc ctgggagctc tctggctaac
     9421 tagggaaccc actgcttaag cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg
     9481 cccgtctgtt gtgtgactct ggtaactaga gatccctcag acccttttag tcagtgtgga
     9541 aaatctctag cagggcccgt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     9601 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     9661 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     9721 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     9781 catgctgggg atgcggtggg ctctatggct tctgaggcgg aaagaaccag ctggggctct
     9841 agggggtatc cccacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg
     9901 cgcagcgtga ccgctacact tgccagcgcc ctagcgcccg ctcctttcgc tttcttccct
     9961 tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta
    10021 gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgatta gggtgatggt
    10081 tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg
    10141 ttctttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
    10201 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
    10261 taacaaaaat ttaacgcgaa ttaattctgt ggaatgtgtg tcagttaggg tgtggaaagt
    10321 ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag tcagcaacca
    10381 ggtgtggaaa gtccccaggc tccccagcag gcagaagtat gcaaagcatg catctcaatt
    10441 agtcagcaac catagtcccg cccctaactc cgcccatccc gcccctaact ccgcccagtt
    10501 ccgcccattc tccgccccat ggctgactaa ttttttttat ttatgcagag gccgaggccg
    10561 cctctgcctc tgagctattc cagaagtagt gaggaggctt ttttggaggc ctaggctttt
    10621 gcaaaaagct cccgggagct tgtatatcca ttttcggatc tgatcagcac gtgttgacaa
    10681 ttaatcatcg gcatagtata tcggcatagt ataatacgac aaggtgagga actaaaccat
    10741 ggccaagttg accagtgccg ttccggtgct caccgcgcgc gacgtcgccg gagcggtcga
    10801 gttctggacc gaccggctcg ggttctcccg ggacttcgtg gaggacgact tcgccggtgt
    10861 ggtccgggac gacgtgaccc tgttcatcag cgcggtccag gaccaggtgg tgccggacaa
    10921 caccctggcc tgggtgtggg tgcgcggcct ggacgagctg tacgccgagt ggtcggaggt
    10981 cgtgtccacg aacttccggg acgcctccgg gccggccatg accgagatcg gcgagcagcc
    11041 gtgggggcgg gagttcgccc tgcgcgaccc ggccggcaac tgcgtgcact tcgtggccga
    11101 ggagcaggac tgacacgtgc tacgagattt cgattccacc gccgccttct atgaaaggtt
    11161 gggcttcgga atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat
    11221 gctggagttc ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag
    11281 caatagcatc acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt
    11341 gtccaaactc atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt
    11401 ggcgtaatca tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca
    11461 caacatacga gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact
    11521 cacattaatt gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct
    11581 gcattaatga atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc
    11641 ttcctcgctc actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca
    11701 ctcaaaggcg gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg
    11761 agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca
    11821 taggctccgc ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa
    11881 cccgacagga ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc
    11941 tgttccgacc ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc
    12001 gctttctcat agctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct
    12061 gggctgtgtg cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg
    12121 tcttgagtcc aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag
    12181 gattagcaga gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta
    12241 cggctacact agaagaacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg
    12301 aaaaagagtt ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt
    12361 tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt
    12421 ttctacgggg tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag
    12481 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
    12541 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
    12601 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
    12661 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
    12721 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
    12781 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
    12841 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
    12901 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
    12961 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
    13021 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
    13081 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
    13141 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
    13201 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
    13261 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
    13321 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
    13381 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
    13441 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt
    13501 tgaatgtatt tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc
    13561 acctgac
//
