LOCUS       sequence-327744-       12948 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  lenti-U6-gRNA::CMVp-nCas9-PmCDA1-UGI-2A-mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 12948)
  AUTHORS   Fukushima T, Tanaka Y, Adachi K, Masuyama N, Tsuchiya A, Asada S, 
            Ishiguro S, Mori H, Seki M, Yachie N, Goyama S, Kitamura T
  TITLE     CRISPR/Cas9-mediated base-editing enables a chain reaction through 
            sequential repair of sgRNA scaffold mutations.
  JOURNAL   Sci Rep. 2021 Dec 13;11(1):23889. doi: 10.1038/s41598-021-02986-6.
  PUBMED    34903756
REFERENCE   2  (bases 1 to 12948)
  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: "Sci Rep."; 
            date: "2021-12-13"; volume: "11(1):23889. doi"; pages: " 
            10.1038/s41598-021-02986-6"
COMMENT     Sequence Label: pLV-CS-154
FEATURES             Location/Qualifiers
     source          1..12948
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     46..65
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(284..388)
                     /gene="bla"
                     /label=AmpR promoter
     polyA_signal    436..570
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(473..492)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     527..546
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     LTR             738..1371
                     /label=3' LTR
                     /note="3' long terminal repeat (LTR) from HIV-1"
     misc_feature    1418..1543
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    2040..2273
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             2458..2502
                     /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    2765..2880
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1 (lacking the first T)"
     promoter        2922..3170
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2922..2942
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     3093..3112
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        3192..3267
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        3336..3639
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3640..3843
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     3794..3814
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             3903..3923
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(3948..3974,3975..3977,3978..8051)
                     /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="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             3948..8051
                     /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
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             8064..8084
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8292..8357
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8991..9011
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             9018..9269
                     /codon_start=1
                     /product="uracil-DNA glycosylase inhibitor from a Bacillus 
                     subtilis bacteriophage (Mol et al., 1995)"
                     /label=UGI
                     /translation="MTNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHT
                     AYDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
     CDS             9270..9323
                     /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             9330..10040
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(9475..9493)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     9743..9762
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     misc_feature    10054..10642
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(10107..10127)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(10525..10536)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             10849..11082
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(11210..11226)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(11210..11226)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(11223..11245)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    11234..11250
                     /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(11258..11264,11265..11282,11283..11288))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11303..11324
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(11441..11458)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(11612..12200)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(11692..11711)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(12371..12948,1..214,215..283))
                     /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"
ORIGIN
        1 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
       61 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
      121 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
      181 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
      241 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
      301 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
      361 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctgacgtcg
      421 acggatcggg agatcaactt gtttattgca gcttataatg gttacaaata aagcaatagc
      481 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
      541 ctcatcaatg tatcttatca tgtctggatc aactggataa ctcaagctaa ccaaaatcat
      601 cccaaacttc ccaccccata ccctattacc actgccaatt acctgtggtt tcatttactc
      661 taaacctgtg attcctctga attattttca ttttaaagaa attgtatttg ttaaatatgt
      721 actacaaact tagtagttgg aagggctaat tcactcccaa agaagacaag atatccttga
      781 tctgtggatc taccacacac aaggctactt ccctgattag cagaactaca caccagggcc
      841 aggggtcaga tatccactga cctttggatg gtgctacaag ctagtaccag ttgagccaga
      901 taaggtagaa gaggccaata aaggagagaa caccagcttg ttacaccctg tgagcctgca
      961 tgggatggat gacccggaga gagaagtgtt agagtggagg tttgacagcc gcctagcatt
     1021 tcatcacgtg gcccgagagc tgcatccgga gtacttcaag aactgctgat atcgagcttg
     1081 ctacaaggga ctttccgctg gggactttcc agggaggcgt ggcctgggcg ggactgggga
     1141 gtggcgagcc ctcagatcct gcatataagc agctgctttt tgcctgtact gggtctctct
     1201 ggttagacca gatctgagcc tgggagctct ctggctaact agggaaccca ctgcttaagc
     1261 ctcaataaag cttgccttga gtgcttcaag tagtgtgtgc ccgtctgttg tgtgactctg
     1321 gtaactagag atccctcaga cccttttagt cagtgtggaa aatctctagc agtggcgccc
     1381 gaacagggac ttgaaagcga aagggaaacc agaggagctc tctcgacgca ggactcggct
     1441 tgctgaagcg cgcacggcaa gaggcgaggg gcggcgactg gtgagtacgc caaaaatttt
     1501 gactagcgga ggctagaagg agagagatgg gtgcgagagc gtcagtatta agcgggggag
     1561 aattagatcg cgatgggaaa aaattcggtt aaggccaggg ggaaagaaaa aatataaatt
     1621 aaaacatata gtatgggcaa gcagggagct agaacgattc gcagttaatc ctggcctgtt
     1681 agaaacatca gaaggctgta gacaaatact gggacagcta caaccatccc ttcagacagg
     1741 atcagaagaa cttagatcat tatataatac agtagcaacc ctctattgtg tgcatcaaag
     1801 gatagagata aaagacacca aggaagcttt agacaagata gaggaagagc aaaacaaaag
     1861 taagaccacc gcacagcaag cggccggccg ctgatcttca gacctggagg aggagatatg
     1921 agggacaatt ggagaagtga attatataaa tataaagtag taaaaattga accattagga
     1981 gtagcaccca ccaaggcaaa gagaagagtg gtgcagagag aaaaaagagc agtgggaata
     2041 ggagctttgt tccttgggtt cttgggagca gcaggaagca ctatgggcgc agcgtcaatg
     2101 acgctgacgg tacaggccag acaattattg tctggtatag tgcagcagca gaacaatttg
     2161 ctgagggcta ttgaggcgca acagcatctg ttgcaactca cagtctgggg catcaagcag
     2221 ctccaggcaa gaatcctggc tgtggaaaga tacctaaagg atcaacagct cctggggatt
     2281 tggggttgct ctggaaaact catttgcacc actgctgtgc cttggaatgc tagttggagt
     2341 aataaatctc tggaacagat ttggaatcac acgacctgga tggagtggga cagagaaatt
     2401 aacaattaca caagcttaat acactcctta attgaagaat cgcaaaacca gcaagaaaag
     2461 aatgaacaag aattattgga attagataaa tgggcaagtt tgtggaattg gtttaacata
     2521 acaaattggc tgtggtatat aaaattattc ataatgatag taggaggctt ggtaggttta
     2581 agaatagttt ttgctgtact ttctatagtg aatagagtta ggcagggata ttcaccatta
     2641 tcgtttcaga cccacctccc aaccccgagg ggacccgaca ggcccgaagg aatagaagaa
     2701 gaaggtggag agagagacag agacagatcc attcgattag tgaacggatc tcgacggtat
     2761 cgcctttaaa agaaaagggg ggattggggg gtacagtgca ggggaaagaa tagtagacat
     2821 aatagcaaca gacatacaaa ctaaagaatt acaaaaacaa attacaaaaa ttcaaaattt
     2881 tcgggtttat tacagggaca gcagagatcc agtttatcga tgagggccta tttcccatga
     2941 ttccttcata tttgcatata cgatacaagg ctgttagaga gataattaga attaatttga
     3001 ctgtaaacac aaagatatta gtacaaaata cgtgacgtag aaagtaataa tttcttgggt
     3061 agtttgcagt tttaaaatta tgttttaaaa tggactatca tatgcttacc gtaacttgaa
     3121 agtatttcga tttcttggct ttatatatct tgtggaaagg acgaaacacc gcacggtcac
     3181 cctgacacgc tgttttagag ctagaaatag caagttaaaa taaggctagt ccgttatcaa
     3241 cttgaaaaag tggcaccgag tcggtgcttt tttactagtt attaatagta atcaattacg
     3301 gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc
     3361 ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc
     3421 atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact
     3481 gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat
     3541 gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact
     3601 tggcagtaca tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac
     3661 atcaatgggc gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac
     3721 gtcaatggga gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac
     3781 tccgccccat tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga
     3841 gctctctggc taactagaga acccactgct tactggcacc ggtaagcgct agcgaaatgg
     3901 caccgaagaa gaagcgtaaa gtcggaatcc acggagttcc tgcggcaatg gacaagaagt
     3961 actccattgg gctcgctatc ggcacaaaca gcgtcggttg ggccgtcatt acggacgagt
     4021 acaaggtgcc gagcaaaaaa ttcaaagttc tgggcaatac cgatcgccac agcataaaga
     4081 agaacctcat tggcgccctc ctgttcgact ccggggagac ggccgaagcc acgcggctca
     4141 aaagaacagc acggcgcaga tatacccgca gaaagaatcg gatctgctac ctgcaggaga
     4201 tctttagtaa tgagatggct aaggtggatg actctttctt ccataggctg gaggagtcct
     4261 ttttggtgga ggaggataaa aagcacgagc gccacccaat ctttggcaat atcgtggacg
     4321 aggtggcgta ccatgaaaag tacccaacca tatatcatct gaggaagaag cttgtagaca
     4381 gtactgataa ggctgacttg cggttgatct atctcgcgct ggcgcatatg atcaaatttc
     4441 ggggacactt cctcatcgag ggggacctga acccagacaa cagcgatgtc gacaaactct
     4501 ttatccaact ggttcagact tacaatcagc ttttcgaaga gaacccgatc aacgcatccg
     4561 gagttgacgc caaagcaatc ctgagcgcta ggctgtccaa atcccggcgg ctcgaaaacc
     4621 tcatcgcaca gctccctggg gagaagaaga acggcctgtt tggtaatctt atcgccctgt
     4681 cactcgggct gacccccaac tttaaatcta acttcgacct ggccgaagat gccaagcttc
     4741 aactgagcaa agacacctac gatgatgatc tcgacaatct gctggcccag atcggcgacc
     4801 agtacgcaga cctttttttg gcggcaaaga acctgtcaga cgccattctg ctgagtgata
     4861 ttctgcgagt gaacacggag atcaccaaag ctccgctgag cgctagtatg atcaagcgct
     4921 atgatgagca ccaccaagac ttgactttgc tgaaggccct tgtcagacag caactgcctg
     4981 agaagtacaa ggaaattttc ttcgatcagt ctaaaaatgg ctacgccgga tacattgacg
     5041 gcggagcaag ccaggaggaa ttttacaaat ttattaagcc catcttggaa aaaatggacg
     5101 gcaccgagga gctgctggta aagcttaaca gagaagatct gttgcgcaaa cagcgcactt
     5161 tcgacaatgg aagcatcccc caccagattc acctgggcga actgcacgct atcctcaggc
     5221 ggcaagagga tttctacccc tttttgaaag ataacaggga aaagattgag aaaatcctca
     5281 catttcggat accctactat gtaggccccc tcgcccgggg aaattccaga ttcgcgtgga
     5341 tgactcgcaa atcagaagag accatcactc cctggaactt cgaggaagtc gtggataagg
     5401 gggcctctgc ccagtccttc atcgaaagga tgactaactt tgataaaaat ctgcctaacg
     5461 aaaaggtgct tcctaaacac tctctgctgt acgagtactt cacagtttat aacgagctca
     5521 ccaaggtcaa atacgtcaca gaagggatga gaaagccagc attcctgtct ggagagcaga
     5581 agaaagctat cgtggacctc ctcttcaaga cgaaccggaa agttaccgtg aaacagctca
     5641 aagaagacta tttcaaaaag attgaatgtt tcgactctgt tgaaatcagc ggagtggagg
     5701 atcgcttcaa cgcatccctg ggaacgtatc acgatctcct gaaaatcatt aaagacaagg
     5761 acttcctgga caatgaggag aacgaggaca ttcttgagga cattgtcctc acccttacgt
     5821 tgtttgaaga tagggagatg attgaagaac gcttgaaaac ttacgctcat ctcttcgacg
     5881 acaaagtcat gaaacagctc aagaggcgcc gatatacagg atgggggcgg ctgtcaagaa
     5941 aactgatcaa tgggatccga gacaagcaga gtggaaagac aatcctggat tttcttaagt
     6001 ccgatggatt tgccaaccgg aacttcatgc agttgatcca tgatgactct ctcaccttta
     6061 aggaggacat ccagaaagca caagtttctg gccaggggga cagtcttcac gagcacatcg
     6121 ctaatcttgc aggtagccca gctatcaaaa agggaatact gcagaccgtt aaggtcgtgg
     6181 atgaactcgt caaagtaatg ggaaggcata agcccgagaa tatcgttatc gagatggccc
     6241 gagagaacca aactacccag aagggacaga agaacagtag ggaaaggatg aagaggattg
     6301 aagagggtat aaaagaactg gggtcccaaa tccttaagga acacccagtt gaaaacaccc
     6361 agcttcagaa tgagaagctc tacctgtact acctgcagaa cggcagggac atgtacgtgg
     6421 atcaggaact ggacatcaat cggctctccg actacgacgt ggatcatatc gtgccccagt
     6481 cttttctcaa agatgattct attgataata aagtgttgac aagatccgat aaaaatagag
     6541 ggaagagtga taacgtcccc tcagaagaag ttgtcaagaa aatgaaaaat tattggcggc
     6601 agctgctgaa cgccaaactg atcacacaac ggaagttcga taatctgact aaggctgaac
     6661 gaggtggcct gtctgagttg gataaagccg gcttcatcaa aaggcagctt gttgagacac
     6721 gccagatcac caagcacgtg gcccaaattc tcgattcacg catgaacacc aagtacgatg
     6781 aaaatgacaa actgattcga gaggtgaaag ttattactct gaagtctaag ctggtctcag
     6841 atttcagaaa ggactttcag ttttataagg tgagagagat caacaattac caccatgcgc
     6901 atgatgccta cctgaatgca gtggtaggca ctgcacttat caaaaaatat cccaagcttg
     6961 aatctgaatt tgtttacgga gactataaag tgtacgatgt taggaaaatg atcgcaaagt
     7021 ctgagcagga aataggcaag gccaccgcta agtacttctt ttacagcaat attatgaatt
     7081 ttttcaagac cgagattaca ctggccaatg gagagattcg gaagcgacca cttatcgaaa
     7141 caaacggaga aacaggagaa atcgtgtggg acaagggtag ggatttcgcg acagtccgga
     7201 aggtcctgtc catgccgcag gtgaacatcg ttaaaaagac cgaagtacag accggaggct
     7261 tctccaagga aagtatcctc ccgaaaagga acagcgacaa gctgatcgca cgcaaaaaag
     7321 attgggaccc caagaaatac ggcggattcg attctcctac agtcgcttac agtgtactgg
     7381 ttgtggccaa agtggagaaa gggaagtcta aaaaactcaa aagcgtcaag gaactgctgg
     7441 gcatcacaat catggagcga tcaagcttcg aaaaaaaccc catcgacttt ctcgaggcga
     7501 aaggatataa agaggtcaaa aaagacctca tcattaagct tcccaagtac tctctctttg
     7561 agcttgaaaa cggccggaaa cgaatgctcg ctagtgcggg cgagctgcag aaaggtaacg
     7621 agctggcact gccctctaaa tacgttaatt tcttgtatct ggccagccac tatgaaaagc
     7681 tcaaagggtc tcccgaagat aatgagcaga agcagctgtt cgtggaacaa cacaaacact
     7741 accttgatga gatcatcgag caaataagcg aattctccaa aagagtgatc ctcgccgacg
     7801 ctaacctcga taaggtgctt tctgcttaca ataagcacag ggataagccc atcagggagc
     7861 aggcagaaaa cattatccac ttgtttactc tgaccaactt gggcgcgcct gcagccttca
     7921 agtacttcga caccaccata gacagaaagc ggtacacctc tacaaaggag gtcctggacg
     7981 ccacactgat tcatcagtca attacggggc tctatgaaac aagaatcgac ctctctcagc
     8041 tcggtggaga cagcagggct gaccccaaga agaagaggaa ggtgggtgga ggaggtaccg
     8101 gcggtggagg ctcagcagaa tacgtacgag ctctgtttga cttcaatggg aatgacgagg
     8161 aggatctccc ctttaagaag ggcgatattc tccgcatcag agataagccc gaagaacaat
     8221 ggtggaatgc cgaggatagc gaagggaaaa ggggcatgat tctggtgcca tatgtggaga
     8281 aatattccgg tgactacaaa gaccatgatg gggattacaa agaccacgac atcgactaca
     8341 aagacgacga cgataaatca gggatgacag acgccgagta cgtgcgcatt catgagaaac
     8401 tggatattta caccttcaag aagcagttct tcaacaacaa gaaatctgtg tcacaccgct
     8461 gctacgtgct gtttgagttg aagcgaaggg gcgaaagaag ggcttgcttt tggggctatg
     8521 ccgtcaacaa gccccaaagt ggcaccgaga gaggaataca cgctgagata ttcagtatcc
     8581 gaaaggtgga agagtatctt cgggataatc ctgggcagtt tacgatcaac tggtattcca
     8641 gctggagtcc ttgcgctgat tgtgccgaga aaattctgga atggtataat caggaacttc
     8701 ggggaaacgg gcacacattg aaaatctggg cctgcaagct gtactacgag aagaatgccc
     8761 ggaaccagat aggactctgg aatctgaggg acaatggtgt aggcctgaac gtgatggttt
     8821 ccgagcacta tcagtgttgt cggaagattt tcatccaaag ctctcataac cagctcaatg
     8881 aaaaccgctg gttggagaaa acactgaaac gtgcggagaa gtggagatcc gagctgagca
     8941 tcatgatcca ggtcaagatt ctgcatacca ctaagtctcc agccgttggt cccaagaaga
     9001 aaagaaaagt cggtaccatg accaaccttt ccgacatcat agagaaggaa acaggcaaac
     9061 agttggtcat ccaagagtcg atactcatgc ttcctgaaga agttgaggag gtcattggga
     9121 ataagccgga aagtgacatt ctcgtacaca ctgcgtatga tgagagcacc gatgagaacg
     9181 tgatgctgct cacgtcagat gccccagagt acaaaccctg ggctctggtg attcaggact
     9241 ctaatggaga gaacaagatc aagatgctag aaggcagggg aagccttctg acttgtgggg
     9301 atgtggaaga aaaccctggt ccatctagaa tggtgagcaa gggcgaggag gataacatgg
     9361 ccatcatcaa ggagttcatg cgcttcaagg tgcacatgga gggctccgtg aacggccacg
     9421 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc
     9481 tgaaggtgac caagggtggc cccctgccct tcgcctggga catcctgtcc cctcagttca
     9541 tgtacggctc caaggcctac gtgaagcacc ccgccgacat ccccgactac ttgaagctgt
     9601 ccttccccga gggcttcaag tgggagcgcg tgatgaactt cgaggacggc ggcgtggtga
     9661 ccgtgaccca ggactcctcc ctgcaggacg gcgagttcat ctacaaggtg aagctgcgcg
     9721 gcaccaactt cccctccgac ggccccgtaa tgcagaagaa gaccatgggc tgggaggcct
     9781 cctccgagcg gatgtacccc gaggacggcg ccctgaaggg cgagatcaag cagaggctga
     9841 agctgaagga cggcggccac tacgacgctg aggtcaagac cacctacaag gccaagaagc
     9901 ccgtgcagct gcccggcgcc tacaacgtca acatcaagtt ggacatcacc tcccacaacg
     9961 aggactacac catcgtggaa cagtacgaac gcgccgaggg ccgccactcc accggcggca
    10021 tggacgagct gtacaagtaa acgcgtctgg aacaatcaac ctctaaatta caaaatttgt
    10081 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
    10141 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
    10201 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
    10261 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
    10321 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
    10381 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
    10441 tcggggaagc tgacgtcctt tccatggctg ctcgcctgtg ttgccacctg gattctgcgc
    10501 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
    10561 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
    10621 tccctttggg ccgcctcccc gcctggaatt aattctgcag tcgagaccta gaaaaacatg
    10681 gagcaatcac aagtagcaat acagcagcta ccaatgctga ttgtgcctgg ctagaagcac
    10741 aagaggagga ggaggtgggt tttccagtca cacctcaggt acctttaaga ccaatgactt
    10801 acaaggcagc tgtagatctt agccactttt taaaagaaaa gaggggactg gaagggctaa
    10861 ttcactccca acgaagacaa gatctgcttt ttgcttgtac tgggtctctc tggttagacc
    10921 agatctgagc ctgggagctc tctggctaac tagggaaccc actgcttaag cctcaataaa
    10981 gcttgccttg agtgcttcaa gtagtgtgtg cccgtctgtt gtgtgactct ggtaactaga
    11041 gatccctcag acccttttag tcagtgtgga aaatctctag cagtagtagt tcatgtcatc
    11101 ttattattca gtatttataa cttgcaaaga aatgaatatc agagagtgag aggccttgac
    11161 attctagcgt tttaccgtcg acctctagct agagcttggc gtaatcatgg tcatagctgt
    11221 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
    11281 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
    11341 tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg
    11401 cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact gactcgctgc
    11461 gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta atacggttat
    11521 ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag caaaaggcca
    11581 ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc
    11641 atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc
    11701 aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg
    11761 gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc tcacgctgta
    11821 ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg
    11881 ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac
    11941 acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag
    12001 gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga agaacagtat
    12061 ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat
    12121 ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc
    12181 gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt
    12241 ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct
    12301 agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt
    12361 ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc
    12421 gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac
    12481 catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat
    12541 cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg
    12601 cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata
    12661 gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta
    12721 tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt
    12781 gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag
    12841 tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa
    12901 gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaat
//
