LOCUS       sequence-413619-       14240 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Lentiviral vector expressing doxycycline-inducible human 
            codon-optimized cytosine base editor where APOBEC1 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 14240)
  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 14240)
  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-APOBEC1
FEATURES             Location/Qualifiers
     source          1..14240
                     /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             863..1546
                     /codon_start=1
                     /product="cytidine deaminase (C to U editing enzyme) from 
                     rat"
                     /label=APOBEC-1
                     /note="can use ssDNA as a substrate (Komor et al., 2016)"
                     /translation="SSETGPVAVDPTLRRRIEPHEFEVFFDPRELRKETCLLYEINWGG
                     RHSIWRHTSQNTNKHVEVNFIEKFTTERYFCPNTRCSITWFLSWSPCGECSRAITEFLS
                     RYPHVTLFIYIARLYHHADPRNRQGLRDLISSGVTIQIMTEQESGYCWRNFVNYSPSNE
                     AHWPRYPHLWVRLYVLELYCIILGLPPCLNILRRKQPQLTFFTIALQSCHYQRLPPHIL
                     WATGLK"
     CDS             join(2087..2110,2111..2113,2114..6187)
                     /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             2087..6187
                     /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             6224..6472
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             6503..6751
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
                     YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             6776..6802
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6812..6838
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6848..6874
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6884..6910
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6920..6946
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             6956..6982
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     regulatory      6992..7001
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     promoter        7001..7212
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     CDS             7228..7623
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     CDS             7630..7683
                     /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(7684..8301,8302..8307,8308..8343,8344..8346,8347..8382,
                     8383..8385,8386..8421,8422..8430)
                     /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    8446..9034
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(8499..8519)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(8917..8928)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             9106..9339
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    9417..9538
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(9454..9473)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     9508..9527
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      9578..9713
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(9733..9752)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(9734..9752)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(9762..9779)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(9762..9778)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9771..9793)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      9920..10375
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(10007..10026)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     10217..10238
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        10401..10505
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(10506..10574,10575..11366)
                     /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(10724..10743)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      11537..12125
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     12026..12045
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     12279..12296
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    12413..12434
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(12449..12454,12455..12472,12473..12479)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    12487..12503
                     /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     12492..12514
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     12511..12527
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     12511..12527
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     12546..12566
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        12548..12566
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        12594..12820
                     /label=RSV promoter
                     /note="Rous sarcoma virus enhancer/promoter"
     LTR             12821..13001
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    13048..13173
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    13666..13899
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             14084..14128
                     /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 cttcctcaga gactgggcct gtcgccgtcg atccaaccct
      901 gcgccgccgg attgaacctc acgagtttga agtgttcttt gacccccggg agctgagaaa
      961 ggagacatgc ctgctgtacg agatcaactg gggaggcagg cactccatct ggaggcacac
     1021 ctctcagaac acaaataagc acgtggaggt gaacttcatc gagaagttta ccacagagcg
     1081 gtacttctgc cccaatacca gatgtagcat cacatggttt ctgagctggt ccccttgcgg
     1141 agagtgtagc agggccatca ccgagttcct gtccagatat ccacacgtga cactgtttat
     1201 ctacatcgcc aggctgtatc accacgcaga cccaaggaat aggcagggcc tgcgcgatct
     1261 gatcagctcc ggcgtgacca tccagatcat gacagagcag gagtccggct actgctggcg
     1321 gaacttcgtg aattattctc ctagcaacga ggcccactgg cctaggtacc cacacctgtg
     1381 ggtgcgcctg tacgtgctgg agctgtattg catcatcctg ggcctgcccc cttgtctgaa
     1441 tatcctgcgg agaaagcagc cccagctgac cttctttaca atcgccctgc agtcttgtca
     1501 ctatcagagg ctgccacccc acatcctgtg ggccacaggc ctgaagagcg gaggatctag
     1561 cggaggatca agcggaagcg agactcctgg aaccagcgaa agcgcaaccc cagaaagcag
     1621 cggaggaagt agcggaggaa gcatggcaat gcagatgcag cttgaagcaa atgcagatac
     1681 ttcagtggaa gaagaaagct ttggcccaca acccatttca cggttagagc agtgtggcat
     1741 aaatgccaac gatgtgaaga aattggaaga agctggattc catactgtgg aggctgttgc
     1801 ctatgcgcca aagaaggagc taataaatat taagggaatt agtgaagcca aagctgataa
     1861 aattctggct gaggcagcta aattagttcc aatgggtttc accactgcaa ctgaattcca
     1921 ccaaaggcgg tcagagatca tacagattac tactggctcc aaagagcttg acaaactact
     1981 tcaatctggc ggatctagcg gtggatctag cggctctgag acccctggaa catccgaatc
     2041 cgccactcca gagagcagcg gaggctcttc tggaggatca agcgctgaca agaagtacag
     2101 catcggcctg gccatcggca ccaactctgt gggctgggcc gtgatcaccg acgagtacaa
     2161 ggtgcccagc aagaaattca aggtgctggg caacaccgac cggcacagca tcaagaagaa
     2221 cctgatcgga gccctgctgt tcgacagcgg cgaaacagcc gaggccaccc ggctgaagag
     2281 aaccgccaga agaagataca ccagacggaa gaaccggatc tgctatctgc aagagatctt
     2341 cagcaacgag atggccaagg tggacgacag cttcttccac agactggaag agtccttcct
     2401 ggtggaagag gataagaagc acgagcggca ccccatcttc ggcaacatcg tggacgaggt
     2461 ggcctaccac gagaagtacc ccaccatcta ccacctgaga aagaaactgg tggacagcac
     2521 cgacaaggcc gacctgcggc tgatctatct ggccctggcc cacatgatca agttccgggg
     2581 ccacttcctg atcgagggcg acctgaaccc cgacaacagc gacgtggaca agctgttcat
     2641 ccagctggtg cagacctaca accagctgtt cgaggaaaac cccatcaacg ccagcggcgt
     2701 ggacgccaag gccatcctgt ctgccagact gagcaagagc agacggctgg aaaatctgat
     2761 cgcccagctg cccggcgaga agaagaatgg cctgttcgga aacctgattg ccctgagcct
     2821 gggcctgacc cccaacttca agagcaactt cgacctggcc gaggatgcca aactgcagct
     2881 gagcaaggac acctacgacg acgacctgga caacctgctg gcccagatcg gcgaccagta
     2941 cgccgacctg tttctggccg ccaagaacct gtccgacgcc atcctgctga gcgacatcct
     3001 gagagtgaac accgagatca ccaaggcccc cctgagcgcc tctatgatca agagatacga
     3061 cgagcaccac caggacctga ccctgctgaa agctctcgtg cggcagcagc tgcctgagaa
     3121 gtacaaagag attttcttcg accagagcaa gaacggctac gccggctaca ttgacggcgg
     3181 agccagccag gaagagttct acaagttcat caagcccatc ctggaaaaga tggacggcac
     3241 cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg cggaagcagc ggaccttcga
     3301 caacggcagc atcccccacc agatccacct gggagagctg cacgccattc tgcggcggca
     3361 ggaagatttt tacccattcc tgaaggacaa ccgggaaaag atcgagaaga tcctgacctt
     3421 ccgcatcccc tactacgtgg gccctctggc caggggaaac agcagattcg cctggatgac
     3481 cagaaagagc gaggaaacca tcaccccctg gaacttcgag gaagtggtgg acaagggcgc
     3541 ttccgcccag agcttcatcg agcggatgac caacttcgat aagaacctgc ccaacgagaa
     3601 ggtgctgccc aagcacagcc tgctgtacga gtacttcacc gtgtataacg agctgaccaa
     3661 agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc ctgagcggcg agcagaaaaa
     3721 ggccatcgtg gacctgctgt tcaagaccaa ccggaaagtg accgtgaagc agctgaaaga
     3781 ggactacttc aagaaaatcg agtgcttcga ctccgtggaa atctccggcg tggaagatcg
     3841 gttcaacgcc tccctgggca cataccacga tctgctgaaa attatcaagg acaaggactt
     3901 cctggacaat gaggaaaacg aggacattct ggaagatatc gtgctgaccc tgacactgtt
     3961 tgaggacaga gagatgatcg aggaacggct gaaaacctat gcccacctgt tcgacgacaa
     4021 agtgatgaag cagctgaagc ggcggagata caccggctgg ggcaggctga gccggaagct
     4081 gatcaacggc atccgggaca agcagtccgg caagacaatc ctggatttcc tgaagtccga
     4141 cggcttcgcc aacagaaact tcatgcagct gatccacgac gacagcctga cctttaaaga
     4201 ggacatccag aaagcccagg tgtccggcca gggcgatagc ctgcacgagc acattgccaa
     4261 tctggccggc agccccgcca ttaagaaggg catcctgcag acagtgaagg tggtggacga
     4321 gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc gtgatcgaaa tggccagaga
     4381 gaaccagacc acccagaagg gacagaagaa cagccgcgag agaatgaagc ggatcgaaga
     4441 gggcatcaaa gagctgggca gccagatcct gaaagaacac cccgtggaaa acacccagct
     4501 gcagaacgag aagctgtacc tgtactacct gcagaatggg cgggatatgt acgtggacca
     4561 ggaactggac atcaaccggc tgtccgacta cgatgtggac catatcgtgc ctcagagctt
     4621 tctgaaggac gactccatcg acaacaaggt gctgaccaga agcgacaaga accggggcaa
     4681 gagcgacaac gtgccctccg aagaggtcgt gaagaagatg aagaactact ggcggcagct
     4741 gctgaacgcc aagctgatta cccagagaaa gttcgacaat ctgaccaagg ccgagagagg
     4801 cggcctgagc gaactggata aggccggctt catcaagaga cagctggtgg aaacccggca
     4861 gatcacaaag cacgtggcac agatcctgga ctcccggatg aacactaagt acgacgagaa
     4921 tgacaagctg atccgggaag tgaaagtgat caccctgaag tccaagctgg tgtccgattt
     4981 ccggaaggat ttccagtttt acaaagtgcg cgagatcaac aactaccacc acgcccacga
     5041 cgcctacctg aacgccgtcg tgggaaccgc cctgatcaaa aagtacccta agctggaaag
     5101 cgagttcgtg tacggcgact acaaggtgta cgacgtgcgg aagatgatcg ccaagagcga
     5161 gcaggaaatc ggcaaggcta ccgccaagta cttcttctac agcaacatca tgaacttttt
     5221 caagaccgag attaccctgg ccaacggcga gatccggaag cggcctctga tcgagacaaa
     5281 cggcgaaacc ggggagatcg tgtgggataa gggccgggat tttgccaccg tgcggaaagt
     5341 gctgagcatg ccccaagtga atatcgtgaa aaagaccgag gtgcagacag gcggcttcag
     5401 caaagagtct atcctgccca agaggaacag cgataagctg atcgccagaa agaaggactg
     5461 ggaccctaag aagtacggcg gcttcgacag ccccaccgtg gcctattctg tgctggtggt
     5521 ggccaaagtg gaaaagggca agtccaagaa actgaagagt gtgaaagagc tgctggggat
     5581 caccatcatg gaaagaagca gcttcgagaa gaatcccatc gactttctgg aagccaaggg
     5641 ctacaaagaa gtgaaaaagg acctgatcat caagctgcct aagtactccc tgttcgagct
     5701 ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa ctgcagaagg gaaacgaact
     5761 ggccctgccc tccaaatatg tgaacttcct gtacctggcc agccactatg agaagctgaa
     5821 gggctccccc gaggataatg agcagaaaca gctgtttgtg gaacagcaca agcactacct
     5881 ggacgagatc atcgagcaga tcagcgagtt ctccaagaga gtgatcctgg ccgacgctaa
     5941 tctggacaaa gtgctgtccg cctacaacaa gcaccgggat aagcccatca gagagcaggc
     6001 cgagaatatc atccacctgt ttaccctgac caatctggga gcccctgccg ccttcaagta
     6061 ctttgacacc accatcgacc ggaagaggta caccagcacc aaagaggtgc tggacgccac
     6121 cctgatccac cagagcatca ccggcctgta cgagacacgg atcgacctgt ctcagctggg
     6181 aggtgacagt ggcggaagtg gactagtagg aagtggcggg agcactaatc tgagcgacat
     6241 cattgagaag gagactggga aacagctggt cattcaggag tccatcctga tgctgcctga
     6301 ggaggtggag gaagtgatcg gcaacaagcc agagtctgac atcctggtgc acaccgccta
     6361 cgacgagtcc acagatgaga atgtgatgct gctgacctct gacgcccccg agtataagcc
     6421 ttgggccctg gtcatccagg attctaacgg cgagaataag atcaagatgc tgagcggagg
     6481 atccggagga tctggaggca gcaccaacct gtctgacatc atcgagaagg agacaggcaa
     6541 gcagctggtc atccaggaga gcatcctgat gctgcccgaa gaagtcgaag aagtgatcgg
     6601 aaacaagcct gagagcgata tcctggtcca taccgcctac gacgagagta ccgacgaaaa
     6661 tgtgatgctg ctgacatccg acgccccaga gtataagccc tgggctctgg tcatccagga
     6721 ttccaacgga gagaacaaaa tcaaaatgct gtctggcggc tcagaattcg gatcccctgc
     6781 tgctaaacgt gttaagcttg atgggggtag cccggcagcc aagagagtca aactcgacgg
     6841 cggctcccca gcggcaaaaa gggtgaaact agacgggggt agccccgccg cgaagcgtgt
     6901 aaagctggat ggaggatcgc ctgcggctaa gcgagtcaaa ttagacggtg ggtcaccggc
     6961 tgcaaagcgc gtcaaattgg attgagctag cgccaccatg gggcagagcg cacatcgccc
     7021 acagtccccg agaagttggg gggaggggtc ggcaattgat ccggtgccta gagaaggtgg
     7081 cgcggggtaa actgggaaag tgatgtcgtg tactggctcc gcctttttcc cgagggtggg
     7141 ggagaaccgt atataagtgc agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc
     7201 gccagaacac agttcgaaca attcaccatg gccaagcctt tgtctcaaga agaatccacc
     7261 ctcattgaaa gagcaacggc tacaatcaac agcatcccca tctctgaaga ctacagcgtc
     7321 gccagcgcag ctctctctag cgacggccgc atcttcactg gtgtcaatgt atatcatttt
     7381 actgggggac cttgtgcaga actcgtggtg ctgggcactg ctgctgctgc ggcagctggc
     7441 aacctgactt gtatcgtcgc gatcggaaat gagaacaggg gcatcttgag cccctgcgga
     7501 cggtgccgac aggtgcttct cgatctgcat cctgggatca aagccatagt gaaggacagt
     7561 gatggacagc cgacggcagt tgggattcgt gaattgctgc cctctggtta tgtgtgggag
     7621 ggcggtaccg aaggtagagg ttctctcctc acttgtggtg atgttgaaga aaaccctggt
     7681 ccaatgtcta gactggacaa gagcaaagtc ataaacggag ctctggaatt actcaatggt
     7741 gtcggtatcg aaggcctgac gacaaggaaa ctcgctcaaa agctgggagt tgagcagcct
     7801 accctgtact ggcacgtgaa gaacaagcgg gccctgctcg atgccctgcc aatcgagatg
     7861 ctggacaggc atcataccca cttctgcccc ctggaaggcg agtcatggca agactttctg
     7921 cggaacaacg ccaagtcata ccgctgtgct ctcctctcac atcgcgacgg ggctaaagtg
     7981 catctcggca cccgcccaac agagaaacag tacgaaaccc tggaaaatca gctcgcgttc
     8041 ctgtgtcagc aaggcttctc cctggagaac gcactgtacg ctctgtccgc cgtgggccac
     8101 tttacactgg gctgcgtatt ggaggaacag gagcatcaag tagcaaaaga ggaaagagag
     8161 acacctacca ccgattctat gcccccactt ctgagacaag caattgagct gttcgaccgg
     8221 cagggagccg aacctgcctt ccttttcggc ctggaactaa tcatatgtgg cctggagaaa
     8281 cagctaaagt gcgaaagcgg cgggccgacc gacgcccttg acgattttga cttagacatg
     8341 ctcccagccg atgcccttga cgactttgac cttgatatgc tgcctgctga cgctcttgac
     8401 gattttgacc ttgacatgct ccccgggtaa acgcgttaag tcgacaatca acctctggat
     8461 tacaaaattt gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt
     8521 ggatacgctg ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc
     8581 tcctccttgt ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg
     8641 caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc
     8701 accacctgtc agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa
     8761 ctcatcgccg cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat
     8821 tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc tgctcgcctg tgttgccacc
     8881 tggattctgc gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt
     8941 ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag
     9001 acgagtcgga tctccctttg ggccgcctcc ccgcgtcgac tttaagacca atgacttaca
     9061 aggcagctgt agatcttagc cactttttaa aagaaaaggg gggactggaa gggctaattc
     9121 actcccaacg aagacaagat ctgctttttg cttgtactgg gtctctctgg ttagaccaga
     9181 tctgagcctg ggagctctct ggctaactag ggaacccact gcttaagcct caataaagct
     9241 tgccttgagt gcttcaagta gtgtgtgccc gtctgttgtg tgactctggt aactagagat
     9301 ccctcagacc cttttagtca gtgtggaaaa tctctagcag tacgtatagt agttcatgtc
     9361 atcttattat tcagtattta taacttgcaa agaaatgaat atcagagagt gagaggaact
     9421 tgtttattgc agcttataat ggttacaaat aaagcaatag catcacaaat ttcacaaata
     9481 aagcattttt ttcactgcat tctagttgtg gtttgtccaa actcatcaat gtatcttatc
     9541 atgtctggct ctagctatcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag
     9601 ttccgcccat tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc
     9661 cgcctcggcc tctgagctat tccagaagta gtgaggaggc ttttttggag gcctagggac
     9721 gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt cgttttacaa
     9781 cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc acatccccct
     9841 ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca acagttgcgc
     9901 agcctgaatg gcgaatggga cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg
     9961 gttacgcgca gcgtgaccgc tacacttgcc agcgccctag cgcccgctcc tttcgctttc
    10021 ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa tcgggggctc
    10081 cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact tgattagggt
    10141 gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt gacgttggag
    10201 tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa ccctatctcg
    10261 gtctattctt ttgatttata agggattttg ccgatttcgg cctattggtt aaaaaatgag
    10321 ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgcttac aatttaggtg
    10381 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
    10441 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
    10501 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
    10561 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
    10621 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
    10681 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
    10741 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
    10801 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
    10861 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
    10921 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
    10981 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
    11041 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
    11101 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
    11161 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
    11221 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
    11281 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
    11341 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
    11401 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
    11461 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
    11521 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
    11581 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
    11641 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
    11701 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
    11761 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
    11821 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
    11881 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
    11941 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
    12001 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
    12061 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
    12121 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
    12181 acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
    12241 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
    12301 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
    12361 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
    12421 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
    12481 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
    12541 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctgcaagct taatgtagtc
    12601 ttatgcaata ctcttgtagt cttgcaacat ggtaacgatg agttagcaac atgccttaca
    12661 aggagagaaa aagcaccgtg catgccgatt ggtggaagta aggtggtacg atcgtgcctt
    12721 attaggaagg caacagacgg gtctgacatg gattggacga accactgaat tgccgcattg
    12781 cagagatatt gtatttaagt gcctagctcg atacataaac gggtctctct ggttagacca
    12841 gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc ctcaataaag
    12901 cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg gtaactagag
    12961 atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc gaacagggac
    13021 ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct tgctgaagcg
    13081 cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt gactagcgga
    13141 ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag aattagatcg
    13201 cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt aaaacatata
    13261 gtatgggcaa gcagggagct agaacgattt gcagttaatc ctggcctgtt agaaacatca
    13321 gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg atcagaagaa
    13381 cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag gatagagata
    13441 aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag taagaccacc
    13501 gcacagcaag cggccgctga tcttcagacc tggaggagga gatatgaggg acaattggag
    13561 aagtgaatta tataaatata aagtagtaaa aattgaacca ttaggagtag cacccaccaa
    13621 ggcaaagaga agagtggtgc agagagaaaa aagagcagtg ggaataggag ctttgttcct
    13681 tgggttcttg ggagcagcag gaagcactat gggcgcagcg tcaatgacgc tgacggtaca
    13741 ggccagacaa ttattgtctg gtatagtgca gcagcagaac aatttgctga gggctattga
    13801 ggcgcaacag catctgttgc aactcacagt ctggggcatc aagcagctcc aggcaagaat
    13861 cctggctgtg gaaagatacc taaaggatca acagctcctg gggatttggg gttgctctgg
    13921 aaaactcatt tgcaccactg ctgtgccttg gaatgctagt tggagtaata aatctctgga
    13981 acagatttgg aatcacacga cctggatgga gtgggacaga gaaattaaca attacacaag
    14041 cttaatacac tccttaattg aagaatcgca aaaccagcaa gaaaagaatg aacaagaatt
    14101 attggaatta gataaatggg caagtttgtg gaattggttt aacataacaa attggctgtg
    14161 gtatataaaa ttattcataa tgatagtagg aggcttggta ggtttaagaa tagtttttgc
    14221 tgtactttct atagtgaata
//
