LOCUS       sequence-181084-       14147 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral plasmid expressing dCas9-2xAM and gRNA cloned in Bbs I 
            sites.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14147)
  AUTHORS   Fujita T, Yuno M, Fujii H
  TITLE     enChIP systems using different CRISPR orthologues and epitope tags.
  JOURNAL   BMC Res Notes. 2018 Feb 27;11(1):154. doi: 
            10.1186/s13104-018-3262-4.
  PUBMED    29482606
REFERENCE   2  (bases 1 to 14147)
  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: "BMC Res 
            Notes."; date: "2018-02-27"; volume: "11(1):154. doi"; pages: " 
            10.1186/s13104-018-3262-4"
COMMENT     Sequence Label: pLenti_dCas9-2xAM
FEATURES             Location/Qualifiers
     source          1..14147
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        616..814
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             832..1012
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    1059..1184
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1676..1909
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2094..2138
                     /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    2412..2529
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2668..2916
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2668..2688
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2839..2858
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2935..3010
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        3107..3392
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        3394..3668
                     /label=chicken beta-actin promoter
     intron          3669..3896
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      3908..3917
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(3914..3940,3941..3943,3944..6427,6428..6433,6434..6499,
                     6500..6502,6503..8017)
                     /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(3914..3940,3941..3943,3944..6430,6431..6433,
                     6434..8017)
                     /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             3914..8017
                     /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             8030..8050
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8156..8212
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             8213..8812
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     complement(8213..8232)
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     primer_bind     8709..8729
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    8842..9430
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8895..8915)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(9313..9324)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9949..10129
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     primer_bind     complement(10155..10172)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    10161..10385
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      10431..10859
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10518..10537)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10728..10749
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(10868..10888)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        10873..11202
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      11053..11188
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     11115..11134
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        11250..11297
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             11316..11690
                     /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    11820..11941
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(11857..11876)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     11911..11930
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(11990..12006)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11990..12006)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(12003..12025)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    12014..12030
                     /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(12038..12044,12045..12062,12063..12068))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12083..12104
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(12221..12238)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(12392..12980)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(12472..12491)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(13151..13942,13943..14011))
                     /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     13774..13793
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(14012..14116)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagcgcgtt ttgcctgtac tgggtctctc
      841 tggttagacc agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag
      901 cctcaataaa gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct
      961 ggtaactaga gatccctcag acccttttag tcagtgtgga aaatctctag cagtggcgcc
     1021 cgaacaggga cttgaaagcg aaagggaaac cagaggagct ctctcgacgc aggactcggc
     1081 ttgctgaagc gcgcacggca agaggcgagg ggcggcgact ggtgagtacg ccaaaaattt
     1141 tgactagcgg aggctagaag gagagagatg ggtgcgagag cgtcagtatt aagcggggga
     1201 gaattagatc gcgatgggaa aaaattcggt taaggccagg gggaaagaaa aaatataaat
     1261 taaaacatat agtatgggca agcagggagc tagaacgatt cgcagttaat cctggcctgt
     1321 tagaaacatc agaaggctgt agacaaatac tgggacagct acaaccatcc cttcagacag
     1381 gatcagaaga acttagatca ttatataata cagtagcaac cctctattgt gtgcatcaaa
     1441 ggatagagat aaaagacacc aaggaagctt tagacaagat agaggaagag caaaacaaaa
     1501 gtaagaccac cgcacagcaa gggccgctga tcttcagacc tggaggagga gatatgaggg
     1561 acaattggag aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag
     1621 cacccaccaa ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag
     1681 ctttgttcct tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc
     1741 tgacggtaca ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga
     1801 gggctattga ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc
     1861 aggcaagaat cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg
     1921 gttgctctgg aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata
     1981 aatctctgga acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca
     2041 attacacaag cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg
     2101 aacaagaatt attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa
     2161 attggctgtg gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa
     2221 tagtttttgc tgtactttct atagtgaata gagttaggca gggatattca ccattatcgt
     2281 ttcagaccca cctcccaacc ccgaggggac ccgacaggcc cgaaggaata gaagaagaag
     2341 gtggagagag agacagagac agatccattc gattagtgaa cggatctaca aatggcagta
     2401 ttcatccaca attttaaaag aaaagggggg attggggggt acagtgcagg ggaaagaata
     2461 gtagacataa tagcaacaga catacaaact aaagaattac aaaaacaaat tacaaaaatt
     2521 caaaattttc gggtttatta cagggacagc agaaattcac tttgaattaa ttcaagcttc
     2581 gtgaggctcc ggtgcccgtc agtgggcaga gcgcacatcg cccacagtcc ccgagaagtt
     2641 ggggggaggg gtcggcaatt gaaccgggag ggcctatttc ccatgattcc ttcatatttg
     2701 catatacgat acaaggctgt tagagagata attggaatta atttgactgt aaacacaaag
     2761 atattagtac aaaatacgtg acgtagaaag taataatttc ttgggtagtt tgcagtttta
     2821 aaattatgtt ttaaaatgga ctatcatatg cttaccgtaa cttgaaagta tttcgatttc
     2881 ttggctttat atatcttgtg gaaaggacga aacaccgggt cttcgagaag acctgtttta
     2941 gagctagaaa tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc
     3001 gagtcggtgc ttttttgttt tagagctaga aatagcaagt taaaataagg ctagtccgtt
     3061 tttagcgcgt gcgccaattc tgcagacaaa tggctctaga ggtacccgtt acataactta
     3121 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaatagtaa
     3181 cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact
     3241 tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta
     3301 aatggcccgc ctggcattgt gcccagtaca tgaccttatg ggactttcct acttggcagt
     3361 acatctacgt attagtcatc gctattacca tggtcgaggt gagccccacg ttctgcttca
     3421 ctctccccat ctcccccccc tccccacccc caattttgta tttatttatt ttttaattat
     3481 tttgtgcagc gatgggggcg gggggggggg ggcgcgcgcc aggcggggcg gggcggggcg
     3541 aggggcgggg cggggcgagg cggagaggtg cggcggcagc caatcagagc ggcgcgctcc
     3601 gaaagtttcc ttttatggcg aggcggcggc ggcggcggcc ctataaaaag cgaagcgcgc
     3661 ggcgggcggg agtcgctgcg cgctgccttc gccccgtgcc ccgctccgcc gccgcctcgc
     3721 gccgcccgcc ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc
     3781 cttctcctcc gggctgtaat tagctgagca agaggtaagg gtttaaggga tggttggttg
     3841 gtggggtatt aatgtttaat tacctggagc acctgcctga aatcactttt tttcaggttg
     3901 gaccggtgcc accatggaca agaagtactc cattgggctc gctatcggca caaacagcgt
     3961 cggctgggcc gtcattacgg acgagtacaa ggtgccgagc aaaaaattca aagttctggg
     4021 caataccgat cgccacagca taaagaagaa cctcattggc gccctcctgt tcgactccgg
     4081 ggagacggcc gaagccacgc ggctcaaaag aacagcacgg cgcagatata cccgcagaaa
     4141 gaatcggatc tgctacctgc aggagatctt tagtaatgag atggctaagg tggatgactc
     4201 tttcttccat aggctggagg agtccttttt ggtggaggag gataaaaagc acgagcgcca
     4261 cccaatcttt ggcaatatcg tggacgaggt ggcgtaccat gaaaagtacc caaccatata
     4321 tcatctgagg aagaagcttg tagacagtac tgataaggct gacttgcggt tgatctatct
     4381 cgcgctggcg catatgatca aatttcgggg acacttcctc atcgaggggg acctgaaccc
     4441 agacaacagc gatgtcgaca aactctttat ccaactggtt cagacttaca atcagctttt
     4501 cgaagagaac ccgatcaacg catccggagt tgacgccaaa gcaatcctga gcgctaggct
     4561 gtccaaatcc cggcggctcg aaaacctcat cgcacagctc cctggggaga agaagaacgg
     4621 cctgtttggt aatcttatcg ccctgtcact cgggctgacc cccaacttta aatctaactt
     4681 cgacctggcc gaagatgcca agcttcaact gagcaaagac acctacgatg atgatctcga
     4741 caatctgctg gcccagatcg gcgaccagta cgcagacctt tttttggcgg caaagaacct
     4801 gtcagacgcc attctgctga gtgatattct gcgagtgaac acggagatca ccaaagctcc
     4861 gctgagcgct agtatgatca agcgctatga tgagcaccac caagacttga ctttgctgaa
     4921 ggcccttgtc agacagcaac tgcctgagaa gtacaaggaa attttcttcg atcagtctaa
     4981 aaatggctac gccggataca ttgacggcgg agcaagccag gaggaatttt acaaatttat
     5041 taagcccatc ttggaaaaaa tggacggcac cgaggagctg ctggtaaagc ttaacagaga
     5101 agatctgttg cgcaaacagc gcactttcga caatggaagc atcccccacc agattcacct
     5161 gggcgaactg cacgctatcc tcaggcggca agaggatttc tacccctttt tgaaagataa
     5221 cagggaaaag attgagaaaa tcctcacatt tcggataccc tactatgtag gccccctcgc
     5281 ccggggaaat tccagattcg cgtggatgac tcgcaaatca gaagagacca tcactccctg
     5341 gaacttcgag gaagtcgtgg ataagggggc ctctgcccag tccttcatcg aaaggatgac
     5401 taactttgat aaaaatctgc ctaacgaaaa ggtgcttcct aaacactctc tgctgtacga
     5461 gtacttcaca gtttataacg agctcaccaa ggtcaaatac gtcacagaag ggatgagaaa
     5521 gccagcattc ctgtctggag agcagaagaa agctatcgtg gacctcctct tcaagacgaa
     5581 ccggaaagtt accgtgaaac agctcaaaga ggactatttc aaaaagattg aatgtttcga
     5641 ctctgttgaa atcagcggag tggaggatcg cttcaacgca tccctgggaa cgtatcacga
     5701 tctcctgaaa atcattaaag acaaggactt cctggacaat gaggagaacg aggacattct
     5761 tgaggacatt gtcctcaccc ttacgttgtt tgaagatagg gagatgattg aagaacgctt
     5821 gaaaacttac gctcatctct tcgacgacaa agtcatgaaa cagctcaaga ggcgccgata
     5881 tacaggatgg gggcggctgt caagaaaact gatcaatggg atcagagaca agcagagtgg
     5941 aaagacaatc ctggattttc ttaagtccga tggatttgcc aacaggaact tcatgcagtt
     6001 gatccatgat gactctctca cctttaagga ggacatccag aaagcacaag tttctggcca
     6061 gggggacagt ctgcacgagc acatcgctaa tcttgcaggt agcccagcta tcaaaaaggg
     6121 aatactgcag accgttaagg tcgtggatga actcgtcaaa gtaatgggaa ggcataagcc
     6181 cgagaatatc gttatcgaga tggcccgaga gaaccaaact acccagaagg gacagaagaa
     6241 cagtagggaa aggatgaaga ggattgaaga gggtataaaa gaactggggt cccaaatcct
     6301 taaggaacac ccagttgaaa acacccagct tcagaatgag aagctctacc tgtactacct
     6361 gcagaacggc agggacatgt acgtggatca ggaactggac atcaatcggc tctccgacta
     6421 cgacgtggat gccatcgtgc cccagtcttt tctcaaagat gattctattg ataataaagt
     6481 gttgacaaga tccgataaaa atagagggaa gagtgataac gtcccctcag aagaagttgt
     6541 caagaaaatg aaaaattatt ggcggcagct gctgaacgcc aaactgatca cacaacggaa
     6601 gttcgataat ctgactaagg ctgaacgagg tggcctgtct gagttggata aagccggctt
     6661 catcaaaagg cagcttgttg agacacgcca gatcaccaag cacgtggccc aaattctcga
     6721 ttcacgcatg aacaccaagt acgatgaaaa tgacaaactg attcgagagg tgaaagttat
     6781 tactctgaag tctaagctgg tctcagattt cagaaaggac tttcagtttt ataaggtgag
     6841 agagatcaac aattaccacc atgcgcatga tgcctacctg aatgcagtgg taggcactgc
     6901 acttatcaaa aaatatccca agcttgaatc tgaatttgtt tacggagact ataaagtgta
     6961 cgatgttagg aaaatgatcg caaagtctga gcaggaaata ggcaaggcca ccgctaagta
     7021 cttcttttac agcaatatta tgaatttttt caagaccgag attacactgg ccaatggaga
     7081 gattcggaag cgaccactta tcgaaacaaa cggagaaaca ggagaaatcg tgtgggacaa
     7141 gggtagggat ttcgcgacag tccggaaggt cctgtccatg ccgcaggtga acatcgttaa
     7201 aaagaccgaa gtacagaccg gaggcttctc caaggaaagt atcctcccga aaaggaacag
     7261 cgacaagctg atcgcacgca aaaaagattg ggaccccaag aaatacggcg gattcgattc
     7321 tcctacagtc gcttacagtg tactggttgt ggccaaagtg gagaaaggga agtctaaaaa
     7381 actcaaaagc gtcaaggaac tgctgggcat cacaatcatg gagcgatcaa gcttcgaaaa
     7441 aaaccccatc gactttctcg aggcgaaagg atataaagag gtcaaaaaag acctcatcat
     7501 taagcttccc aagtactctc tctttgagct tgaaaacggc cggaaacgaa tgctcgctag
     7561 tgcgggcgag ctgcagaaag gtaacgagct ggcactgccc tctaaatacg ttaatttctt
     7621 gtatctggcc agccactatg aaaagctcaa agggtctccc gaagataatg agcagaagca
     7681 gctgttcgtg gaacaacaca aacactacct tgatgagatc atcgagcaaa taagcgaatt
     7741 ctccaaaaga gtgatcctcg ccgacgctaa cctcgataag gtgctttctg cttacaataa
     7801 gcacagggat aagcccatca gggagcaggc agaaaacatt atccacttgt ttactctgac
     7861 caacttgggc gcgcctgcag ccttcaagta cttcgacacc accatagaca gaaagcggta
     7921 cacctctaca aaggaggtcc tggacgccac actgattcat cagtcaatta cggggctcta
     7981 tgaaacaaga atcgacctct ctcagctcgg tggagacagc agggctgacc ccaagaagaa
     8041 gaggaaggtg gggcaagatc ctcaacgcaa aggcaacgtg atactctctc aggcttacgg
     8101 gcaagatcct caacgcaaag gcaacgtgat actctctcag gcttacggat ccggagctac
     8161 taacttcagc ctgctgaagc aggctggaga cgtggaggag aaccctggac ctatgaccga
     8221 gtacaagccc acggtgcgcc tcgccacccg cgacgacgtc cccagggccg tacgcaccct
     8281 cgccgccgcg ttcgccgact accccgccac gcgccacacc gtcgatccgg accgccacat
     8341 cgagcgggtc accgagctgc aagaactctt cctcacgcgc gtcgggctcg acatcggcaa
     8401 ggtgtgggtc gcggacgacg gcgccgcggt ggcggtctgg accacgccgg agagcgtcga
     8461 agcgggggcg gtgttcgccg agatcggccc gcgcatggcc gagttgagcg gttcccggct
     8521 ggccgcgcag caacagatgg aaggcctcct ggcgccgcac cggcccaagg agcccgcgtg
     8581 gttcctggcc accgtcggcg tctcgcccga ccaccagggc aagggtctgg gcagcgccgt
     8641 cgtgctcccc ggagtggagg cggccgagcg cgccggggtg cccgccttcc tggagacctc
     8701 cgcgccccgc aacctcccct tctacgagcg gctcggcttc accgtcaccg ccgacgtcga
     8761 ggtgcccgaa ggaccgcgca cctggtgcat gacccgcaag cccggtgcct gagttaacat
     8821 cgagggatca agcttatcga taatcaacct ctggattaca aaatttgtga aagattgact
     8881 ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt aatgcctttg
     8941 tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa atcctggttg
     9001 ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt gtgcactgtg
     9061 tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct cctttccggg
     9121 actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg ccttgcccgc
     9181 tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc ggggaagctg
     9241 acgtcctttc catggctgct cgcctgtgtt gccacctgga ttctgcgcgg gacgtccttc
     9301 tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct gctgccggct
     9361 ctgcggcctc ttccgcgtat tcgccttcgc cctcagacga gtcggatctc cctttgggcc
     9421 gcctccccgc atcgataccg tcgagaccta gaaaaacatg gagcaatcac aagtagcaat
     9481 acagcagcta ccaatgctga ttgtgcctgg ctagaagcac aagaggagga ggaggtgggt
     9541 tttccagtca cacctcaggt acctttaaga ccaatgactt acaaggcagc tgtagatctt
     9601 agccactttt taaaagaaaa ggggggactg gaagggctaa ttcactccca acgatgacaa
     9661 gatatccttg atctgtggat ctaccacaca caaggctact tccctgattg gcagaactac
     9721 acaccagggc cagggatcag atatccactg acctttggat ggtgctacaa gctagtacca
     9781 gttgagcaag agaaggtaga agaagccaat gaaggagaga acacccgctt gttacaccct
     9841 gtgagcctgc atgggatgga tgacccggag agagaagtat tagagtggag gtttgacagc
     9901 cgcctagcat ttcatcacat ggcccgagag ctgcatccgg actgtactgg gtctctctgg
     9961 ttagaccaga tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct
    10021 caataaagct tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt
    10081 aactagagat ccctcagacc cttttagtca gtgtggaaaa tctctagcag ggcccgttta
    10141 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
    10201 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
    10261 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
    10321 ggacagcaag ggggaggatt gggatgacaa tagcaggcat gctggggatg cggtgggctc
    10381 tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc acgcgccctg
    10441 tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc
    10501 cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg
    10561 ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta gtgctttacg
    10621 gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc catcgccctg
    10681 atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg gactcttgtt
    10741 ccaaactgga acaacactca accctatctc ggtctattct tttgatttat aagggatttt
    10801 gccgatttcg gcctattggt taaaaaatga gctgatttaa caaaaattta acgcgaatta
    10861 attctgtgga atgtgtgtca gttagggtgt ggaaagtccc caggctcccc agcaggcaga
    10921 agtatgcaaa gcatgcatct caattagtca gcaaccaggt gtggaaagtc cccaggctcc
    10981 ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccat agtcccgccc
    11041 ctaactccgc ccatcccgcc cctaactccg cccagttccg cccattctcc gccccatggc
    11101 tgactaattt tttttattta tgcagaggcc gaggccgcct ctgcctctga gctattccag
    11161 aagtagtgag gaggcttttt tggaggccta ggcttttgca aaaagctccc gggagcttgt
    11221 atatccattt tcggatctga tcagcacgtg ttgacaatta atcatcggca tagtatatcg
    11281 gcatagtata atacgacaag gtgaggaact aaaccatggc caagttgacc agtgccgttc
    11341 cggtgctcac cgcgcgcgac gtcgccggag cggtcgagtt ctggaccgac cggctcgggt
    11401 tctcccggga cttcgtggag gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt
    11461 tcatcagcgc ggtccaggac caggtggtgc cggacaacac cctggcctgg gtgtgggtgc
    11521 gcggcctgga cgagctgtac gccgagtggt cggaggtcgt gtccacgaac ttccgggacg
    11581 cctccgggcc ggccatgacc gagatcggcg agcagccgtg ggggcgggag ttcgccctgc
    11641 gcgacccggc cggcaactgc gtgcacttcg tggccgagga gcaggactga cacgtgctac
    11701 gagatttcga ttccaccgcc gccttctatg aaaggttggg cttcggaatc gttttccggg
    11761 acgccggctg gatgatcctc cagcgcgggg atctcatgct ggagttcttc gcccacccca
    11821 acttgtttat tgcagcttat aatggttaca aataaagcaa tagcatcaca aatttcacaa
    11881 ataaagcatt tttttcactg cattctagtt gtggtttgtc caaactcatc aatgtatctt
    11941 atcatgtctg tataccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
    12001 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
    12061 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
    12121 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
    12181 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
    12241 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
    12301 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
    12361 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
    12421 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
    12481 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
    12541 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
    12601 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
    12661 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
    12721 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
    12781 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
    12841 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
    12901 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
    12961 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
    13021 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
    13081 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
    13141 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
    13201 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
    13261 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
    13321 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
    13381 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
    13441 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
    13501 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
    13561 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
    13621 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
    13681 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc
    13741 gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt
    13801 taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc
    13861 tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta
    13921 ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa
    13981 taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca
    14041 tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac
    14101 aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
