LOCUS       sequence-413592-       14150 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector expressing doxycycline-inducible human 
            codon-optimized cytosine base editor where APOBEC3A is fused to the 
            N-terminus of SpCas9(D10A)-sDNA Rad51-2xUGI-6xNLS .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14150)
  AUTHORS   Xiao MS, Damodaran AP, Kumari B, Dickson E, Xing K, On TA, Parab N, 
            King HE, Perez AR, Guiblet WM, Duncan G, Che A, Chari R, Andresson 
            T, Vidigal JA, Weatheritt RJ, Aregger M, Gonatopoulos-Pournatzis T
  TITLE     Genome-scale exon perturbation screens uncover exons critical for 
            cell fitness.
  JOURNAL   Mol Cell. 2024 Jul 11;84(13):2553-2572.e19. doi: 
            10.1016/j.molcel.2024.05.024. Epub 2024 Jun 24.
  PUBMED    38917794
REFERENCE   2  (bases 1 to 14150)
  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"; doi: 
            "10.1016/j.molcel.2024.05.024"; journalName: "Mol Cell"; date: 
            "2024-07-11- 11"; volume: "84"; issue: "13"; pages: "2553-2572.e19"
COMMENT     Sequence Label: pLenti-nSpCas9-APOBEC3A
FEATURES             Location/Qualifiers
     source          1..14150
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     66..86
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     misc_feature    287..404
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        469..783
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    477..495
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    513..531
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    548..566
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    584..602
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    620..638
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    655..673
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    691..709
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     756..780
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             806..829
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             830..856
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             join(1997..2020,2021..2023,2024..6097)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1997..6097
                     /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="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             6134..6382
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             6413..6661
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             6686..6712
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6722..6748
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6758..6784
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6794..6820
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6830..6856
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6866..6892
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     regulatory      6902..6911
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        6911..7122
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             7138..7533
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             7540..7593
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(7594..8211,8212..8217,8218..8253,8254..8256,8257..8292,
                     8293..8295,8296..8331,8332..8340)
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSYRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPTDALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     misc_feature    8356..8944
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8409..8429)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8827..8838)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9016..9249
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    9327..9448
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(9364..9383)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     9418..9437
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      9488..9623
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(9643..9662)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(9644..9662)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9672..9689)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(9672..9688)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9681..9703)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9830..10285
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(9917..9936)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10127..10148
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        10311..10415
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10416..10484,10485..11276)
                     /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     complement(10634..10653)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11447..12035
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     11936..11955
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     12189..12206
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    12323..12344
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(12359..12364,12365..12382,12383..12389)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12397..12413
                     /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)."
     primer_bind     12402..12424
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     12421..12437
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     12421..12437
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     12456..12476
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        12458..12476
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        12504..12730
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             12731..12911
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    12958..13083
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    13576..13809
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             13994..14038
                     /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"
ORIGIN
        1 gagttaggca gggatattca ccattatcgt ttcagaccca cctcccaacc ccgaggggac
       61 ccagagaggg cctatttccc atgattcctt catatttgca tatacgatac aaggctgtta
      121 gagagataat tagaattaat ttgactgtaa acacaaagat attagtacaa aatacgtgac
      181 gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt aaaatggact
      241 atcataaatt ctagatcttg agacaaatgg cagtattcat ccacaatttt aaaagaaaag
      301 gggggattgg ggggtacagt gcaggggaaa gaatagtaga cataatagca acagacatac
      361 aaactaaaga attacaaaaa caaattacaa aaattcaaaa ttttcgggtt tattacaggg
      421 acagcagaga tccactttgg cgccggctcg agcgcgaatc gatatgtcga gtttactccc
      481 tatcagtgat agagaacgta tgtcgagttt actccctatc agtgatagag aacgatgtcg
      541 agtttactcc ctatcagtga tagagaacgt atgtcgagtt tactccctat cagtgataga
      601 gaacgtatgt cgagtttact ccctatcagt gatagagaac gtatgtcgag tttatcccta
      661 tcagtgatag agaacgtatg tcgagtttac tccctatcag tgatagagaa cgtatgtcga
      721 ggtaggcgtg tacggtggga ggcctatata agcagagctc gtttagtgaa ccgtcagatc
      781 gcctggagaa ttggctagca ccatggatta caaagacgat gacgataagt acccatacga
      841 tgtgccagat tacgccagcg ctgaggcatc tccagcaagc ggaccaaggc acctgatgga
      901 cccccacatc ttcacctcta actttaacaa tggcatcggc aggcacaaga catacctgtg
      961 ctatgaggtg gagcgcctgg acaacggcac cagcgtgaag atggatcagc acagaggctt
     1021 cctgcacaac caggccaaga atctgctgtg cggcttctac ggccggcacg cagagctgag
     1081 atttctggac ctggtgccta gcctgcagct ggatccagcc cagatctata gggtgacctg
     1141 gttcatcagc tggtccccat gcttttcctg gggatgtgca ggagaggtgc gcgccttcct
     1201 gcaggagaat acacacgtgc ggctgagaat ctttgccgcc cggatctacg actatgatcc
     1261 tctgtacaag gaggccctgc agatgctgag agacgcagga gcccaggtgt ccatcatgac
     1321 ctatgatgag ttcaagcact gctgggacac atttgtggat caccagggct gtccctttta
     1381 gccttgggac ggactggatg agcactccca ggccctgtct ggcaggctga gggccatcct
     1441 gcagaaccag ggcaatagcg gaggatctag cggaggatca agcggaagcg agactcctgg
     1501 aaccagcgaa agcgcaaccc cagaaagcag cggaggaagt agcggaggaa gcatggcaat
     1561 gcagatgcag cttgaagcaa atgcagatac ttcagtggaa gaagaaagct ttggcccaca
     1621 acccatttca cggttagagc agtgtggcat aaatgccaac gatgtgaaga aattggaaga
     1681 agctggattc catactgtgg aggctgttgc ctatgcgcca aagaaggagc taataaatat
     1741 taagggaatt agtgaagcca aagctgataa aattctggct gaggcagcta aattagttcc
     1801 aatgggtttc accactgcaa ctgaattcca ccaaaggcgg tcagagatca tacagattac
     1861 tactggctcc aaagagcttg acaaactact tcaatctggc ggatctagcg gtggatctag
     1921 cggctctgag acccctggaa catccgaatc cgccactcca gagagcagcg gaggctcttc
     1981 tggaggatca agcgctgaca agaagtacag catcggcctg gccatcggca ccaactctgt
     2041 gggctgggcc gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg
     2101 caacaccgac cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg
     2161 cgaaacagcc gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa
     2221 gaaccggatc tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag
     2281 cttcttccac agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca
     2341 ccccatcttc ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta
     2401 ccacctgaga aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct
     2461 ggccctggcc cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc
     2521 cgacaacagc gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt
     2581 cgaggaaaac cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact
     2641 gagcaagagc agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg
     2701 cctgttcgga aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt
     2761 cgacctggcc gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga
     2821 caacctgctg gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct
     2881 gtccgacgcc atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc
     2941 cctgagcgcc tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa
     3001 agctctcgtg cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa
     3061 gaacggctac gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat
     3121 caagcccatc ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga
     3181 ggacctgctg cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct
     3241 gggagagctg cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa
     3301 ccgggaaaag atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc
     3361 caggggaaac agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg
     3421 gaacttcgag gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac
     3481 caacttcgat aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga
     3541 gtacttcacc gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa
     3601 gcccgccttc ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa
     3661 ccggaaagtg accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcttcga
     3721 ctccgtggaa atctccggcg tggaagatcg gttcaacgcc tccctgggca cataccacga
     3781 tctgctgaaa attatcaagg acaaggactt cctggacaat gaggaaaacg aggacattct
     3841 ggaagatatc gtgctgaccc tgacactgtt tgaggacaga gagatgatcg aggaacggct
     3901 gaaaacctat gcccacctgt tcgacgacaa agtgatgaag cagctgaagc ggcggagata
     3961 caccggctgg ggcaggctga gccggaagct gatcaacggc atccgggaca agcagtccgg
     4021 caagacaatc ctggatttcc tgaagtccga cggcttcgcc aacagaaact tcatgcagct
     4081 gatccacgac gacagcctga cctttaaaga ggacatccag aaagcccagg tgtccggcca
     4141 gggcgatagc ctgcacgagc acattgccaa tctggccggc agccccgcca ttaagaaggg
     4201 catcctgcag acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc
     4261 cgagaacatc gtgatcgaaa tggccagaga gaaccagacc acccagaagg gacagaagaa
     4321 cagccgcgag agaatgaagc ggatcgaaga gggcatcaaa gagctgggca gccagatcct
     4381 gaaagaacac cccgtggaaa acacccagct gcagaacgag aagctgtacc tgtactacct
     4441 gcagaatggg cgggatatgt acgtggacca ggaactggac atcaaccggc tgtccgacta
     4501 cgatgtggac catatcgtgc ctcagagctt tctgaaggac gactccatcg acaacaaggt
     4561 gctgaccaga agcgacaaga accggggcaa gagcgacaac gtgccctccg aagaggtcgt
     4621 gaagaagatg aagaactact ggcggcagct gctgaacgcc aagctgatta cccagagaaa
     4681 gttcgacaat ctgaccaagg ccgagagagg cggcctgagc gaactggata aggccggctt
     4741 catcaagaga cagctggtgg aaacccggca gatcacaaag cacgtggcac agatcctgga
     4801 ctcccggatg aacactaagt acgacgagaa tgacaagctg atccgggaag tgaaagtgat
     4861 caccctgaag tccaagctgg tgtccgattt ccggaaggat ttccagtttt acaaagtgcg
     4921 cgagatcaac aactaccacc acgcccacga cgcctacctg aacgccgtcg tgggaaccgc
     4981 cctgatcaaa aagtacccta agctggaaag cgagttcgtg tacggcgact acaaggtgta
     5041 cgacgtgcgg aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta
     5101 cttcttctac agcaacatca tgaacttttt caagaccgag attaccctgg ccaacggcga
     5161 gatccggaag cggcctctga tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa
     5221 gggccgggat tttgccaccg tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa
     5281 aaagaccgag gtgcagacag gcggcttcag caaagagtct atcctgccca agaggaacag
     5341 cgataagctg atcgccagaa agaaggactg ggaccctaag aagtacggcg gcttcgacag
     5401 ccccaccgtg gcctattctg tgctggtggt ggccaaagtg gaaaagggca agtccaagaa
     5461 actgaagagt gtgaaagagc tgctggggat caccatcatg gaaagaagca gcttcgagaa
     5521 gaatcccatc gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat
     5581 caagctgcct aagtactccc tgttcgagct ggaaaacggc cggaagagaa tgctggcctc
     5641 tgccggcgaa ctgcagaagg gaaacgaact ggccctgccc tccaaatatg tgaacttcct
     5701 gtacctggcc agccactatg agaagctgaa gggctccccc gaggataatg agcagaaaca
     5761 gctgtttgtg gaacagcaca agcactacct ggacgagatc atcgagcaga tcagcgagtt
     5821 ctccaagaga gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa
     5881 gcaccgggat aagcccatca gagagcaggc cgagaatatc atccacctgt ttaccctgac
     5941 caatttggga gcccctgccg cctttaagta ctttgacacc accatcgacc ggaagaggta
     6001 caccagcacc aaagaggtgc tggacgccac cctgatccac cagagcatca ccggcctgta
     6061 cgagacacgg atcgacctgt ctcagctggg aggtgacagt ggcggaagtg gactagtagg
     6121 aagtggcggg agcactaatc tgagcgacat cattgagaag gagactggga aacagctggt
     6181 cattcaggag tccatcctga tgctgcctga ggaggtggag gaagtgatcg gcaacaagcc
     6241 agagtctgac atcctggtgc acaccgccta cgacgagtcc acagatgaga atgtgatgct
     6301 gctgacctct gacgcccccg agtataagcc ttgggccctg gtcatccagg attctaacgg
     6361 cgagaataag atcaagatgc tgagcggagg atccggagga tctggaggca gcaccaacct
     6421 gtctgacatc atcgagaagg agacaggcaa gcagctggtc atccaggaga gcatcctgat
     6481 gctgcccgaa gaagtcgaag aagtgatcgg aaacaagcct gagagcgata tcctggtcca
     6541 taccgcctac gacgagagta ccgacgaaaa tgtgatgctg ctgacatccg acgccccaga
     6601 gtataagccc tgggctctgg tcatccagga ttccaacgga gagaacaaaa tcaaaatgct
     6661 gtctggcggc tcagaattcg gatcccctgc tgctaaacgt gttaagcttg atgggggtag
     6721 cccggcagcc aagagagtca aactcgacgg cggctcccca gcggcaaaaa gggtgaaact
     6781 agacgggggt agccccgccg cgaagcgtgt aaagctggat ggaggatcgc ctgcggctaa
     6841 gcgagtcaaa ttagacggtg ggtcaccggc tgcaaagcgc gtcaaattgg attgagctag
     6901 cgccaccatg gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc
     6961 ggcaattgat ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg
     7021 tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc
     7081 gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac agttcgaaca attcaccatg
     7141 gccaagcctt tgtctcaaga agaatccacc ctcattgaaa gagcaacggc tacaatcaac
     7201 agcatcccca tctctgaaga ctacagcgtc gccagcgcag ctctctctag cgacggccgc
     7261 atcttcactg gtgtcaatgt atatcatttt actgggggac cttgtgcaga actcgtggtg
     7321 ctgggcactg ctgctgctgc ggcagctggc aacctgactt gtatcgtcgc gatcggaaat
     7381 gagaacaggg gcatcttgag cccctgcgga cggtgccgac aggtgcttct cgatctgcat
     7441 cctgggatca aagccatagt gaaggacagt gatggacagc cgacggcagt tgggattcgt
     7501 gaattgctgc cctctggtta tgtgtgggag ggcggtaccg aaggtagagg ttctctcctc
     7561 acttgtggtg atgttgaaga aaaccctggt ccaatgtcta gactggacaa gagcaaagtc
     7621 ataaacggag ctctggaatt actcaatggt gtcggtatcg aaggcctgac gacaaggaaa
     7681 ctcgctcaaa agctgggagt tgagcagcct accctgtact ggcacgtgaa gaacaagcgg
     7741 gccctgctcg atgccctgcc aatcgagatg ctggacaggc atcataccca cttctgcccc
     7801 ctggaaggcg agtcatggca agactttctg cggaacaacg ccaagtcata ccgctgtgct
     7861 ctcctctcac atcgcgacgg ggctaaagtg catctcggca cccgcccaac agagaaacag
     7921 tacgaaaccc tggaaaatca gctcgcgttc ctgtgtcagc aaggcttctc cctggagaac
     7981 gcactgtacg ctctgtccgc cgtgggccac tttacactgg gctgcgtatt ggaggaacag
     8041 gagcatcaag tagcaaaaga ggaaagagag acacctacca ccgattctat gcccccactt
     8101 ctgagacaag caattgagct gttcgaccgg cagggagccg aacctgcctt ccttttcggc
     8161 ctggaactaa tcatatgtgg cctggagaaa cagctaaagt gcgaaagcgg cgggccgacc
     8221 gacgcccttg acgattttga cttagacatg ctcccagccg atgcccttga cgactttgac
     8281 cttgatatgc tgcctgctga cgctcttgac gattttgacc ttgacatgct ccccgggtaa
     8341 acgcgttaag tcgacaatca acctctggat tacaaaattt gtgaaagatt gactggtatt
     8401 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
     8461 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
     8521 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
     8581 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
     8641 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
     8701 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
     8761 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
     8821 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
     8881 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
     8941 ccgcgtcgac tttaagacca atgacttaca aggcagctgt agatcttagc cactttttaa
     9001 aagaaaaggg gggactggaa gggctaattc actcccaacg aagacaagat ctgctttttg
     9061 cttgtactgg gtctctctgg ttagaccaga tctgagcctg ggagctctct ggctaactag
     9121 ggaacccact gcttaagcct caataaagct tgccttgagt gcttcaagta gtgtgtgccc
     9181 gtctgttgtg tgactctggt aactagagat ccctcagacc cttttagtca gtgtggaaaa
     9241 tctctagcag tacgtatagt agttcatgtc atcttattat tcagtattta taacttgcaa
     9301 agaaatgaat atcagagagt gagaggaact tgtttattgc agcttataat ggttacaaat
     9361 aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg
     9421 gtttgtccaa actcatcaat gtatcttatc atgtctggct ctagctatcc cgcccctaac
     9481 tccgcccatc ccgcccctaa ctccgcccag ttccgcccat tctccgcccc atggctgact
     9541 aatttttttt atttatgcag aggccgaggc cgcctcggcc tctgagctat tccagaagta
     9601 gtgaggaggc ttttttggag gcctagggac gtacccaatt cgccctatag tgagtcgtat
     9661 tacgcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     9721 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     9781 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggga cgcgccctgt
     9841 agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc
     9901 agcgccctag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc
     9961 tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg
    10021 cacctcgacc ccaaaaaact tgattagggt gatggttcac gtagtgggcc atcgccctga
    10081 tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg actcttgttc
    10141 caaactggaa caacactcaa ccctatctcg gtctattctt ttgatttata agggattttg
    10201 ccgatttcgg cctattggtt aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt
    10261 aacaaaatat taacgcttac aatttaggtg gcacttttcg gggaaatgtg cgcggaaccc
    10321 ctatttgttt atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct
    10381 gataaatgct tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg
    10441 cccttattcc cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg
    10501 tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc
    10561 tcaacagcgg taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca
    10621 cttttaaagt tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac
    10681 tcggtcgccg catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa
    10741 agcatcttac ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg
    10801 ataacactgc ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt
    10861 ttttgcacaa catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg
    10921 aagccatacc aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc
    10981 gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga
    11041 tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta
    11101 ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc
    11161 cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg
    11221 atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt
    11281 cagaccaagt ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa
    11341 ggatctaggt gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt
    11401 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
    11461 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
    11521 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
    11581 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
    11641 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
    11701 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
    11761 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
    11821 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
    11881 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
    11941 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
    12001 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
    12061 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
    12121 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
    12181 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
    12241 tccccgcgcg ttggccgatt cattaatgca gctggcacga caggtttccc gactggaaag
    12301 cgggcagtaa gcgcaacgca attaatgtga gttagctcac tcattaggca ccccaggctt
    12361 tacactttat gcttccggct cgtatgttgt gtggaattgt gagcggataa caatttcaca
    12421 caggaaacag ctatgaccat gattacgcca agcgcgcaat taaccctcac taaagggaac
    12481 aaaagctgga gctgcaagct taatgtagtc ttatgcaata ctcttgtagt cttgcaacat
    12541 ggtaacgatg agttagcaac atgccttaca aggagagaaa aagcaccgtg catgccgatt
    12601 ggtggaagta aggtggtacg atcgtgcctt attaggaagg caacagacgg gtctgacatg
    12661 gattggacga accactgaat tgccgcattg cagagatatt gtatttaagt gcctagctcg
    12721 atacataaac gggtctctct ggttagacca gatctgagcc tgggagctct ctggctaact
    12781 agggaaccca ctgcttaagc ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc
    12841 ccgtctgttg tgtgactctg gtaactagag atccctcaga cccttttagt cagtgtggaa
    12901 aatctctagc agtggcgccc gaacagggac ttgaaagcga aagggaaacc agaggagctc
    12961 tctcgacgca ggactcggct tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg
    13021 gtgagtacgc caaaaatttt gactagcgga ggctagaagg agagagatgg gtgcgagagc
    13081 gtcagtatta agcgggggag aattagatcg cgatgggaaa aaattcggtt aaggccaggg
    13141 ggaaagaaaa aatataaatt aaaacatata gtatgggcaa gcagggagct agaacgattc
    13201 gcagttaatc ctggcctgtt agaaacatca gaaggctgta gacaaatact gggacagcta
    13261 caaccatccc ttcagacagg atcagaagaa cttagatcat tatataatac agtagcaacc
    13321 ctctattgtg tgcatcaaag gatagagata aaagacacca aggaagcttt agacaagata
    13381 gaggaagagc aaaacaaaag taagaccacc gcacagcaag cggccgctga tcttcagacc
    13441 tggaggagga gatatgaggg acaattggag aagtgaatta tataaatata aagtagtaaa
    13501 aattgaacca ttaggagtag cacccaccaa ggcaaagaga agagtggtgc agagagaaaa
    13561 aagagcagtg ggaataggag ctttgttcct tgggttcttg ggagcagcag gaagcactat
    13621 gggcgcagcg tcaatgacgc tgacggtaca ggccagacaa ttattgtctg gtatagtgca
    13681 gcagcagaac aatttgctga gggctattga ggcgcaacag catctgttgc aactcacagt
    13741 ctggggcatc aagcagctcc aggcaagaat cctggctgtg gaaagatacc taaaggatca
    13801 acagctcctg gggatttggg gttgctctgg aaaactcatt tgcaccactg ctgtgccttg
    13861 gaatgctagt tggagtaata aatctctgga acagatttgg aatcacacga cctggatgga
    13921 gtgggacaga gaaattaaca attacacaag cttaatacac tccttaattg aagaatcgca
    13981 aaaccagcaa gaaaagaatg aacaagaatt attggaatta gataaatggg caagtttgtg
    14041 gaattggttt aacataacaa attggctgtg gtatataaaa ttattcataa tgatagtagg
    14101 aggcttggta ggtttaagaa tagtttttgc tgtactttct atagtgaata
//
