LOCUS       sequence-334780-       13013 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9 and guide RNA for ACTB gene in human.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13013)
  AUTHORS   Yoshino Y, Endo S, Chen Z, Qi H, Watanabe G, Chiba N
  TITLE     Evaluation of site-specific homologous recombination activity of 
            BRCA1 by direct quantitation of gene editing efficiency.
  JOURNAL   Sci Rep. 2019 Feb 7;9(1):1644. doi: 10.1038/s41598-018-38311-x.
  PUBMED    30733539
REFERENCE   2  (bases 1 to 13013)
  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: "2019-02-7"; volume: "9(1):1644. doi"; pages: " 
            10.1038/s41598-018-38311-x"
COMMENT     Sequence Label: LentiCRISPRv2-ACTB-C1
FEATURES             Location/Qualifiers
     source          1..13013
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        27..225
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     180..200
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     LTR             243..423
                     /label=5' LTR (truncated)
                     /note="truncated 5' long terminal repeat (LTR) from HIV-1"
     misc_feature    470..595
                     /label=HIV-1 Psi
                     /note="packaging signal of human immunodeficiency virus 
                     type 1"
     misc_feature    1088..1321
                     /label=RRE
                     /note="The Rev response element (RRE) of HIV-1 allows for 
                     Rev-dependent mRNA export from the nucleus to the 
                     cytoplasm."
     CDS             1506..1550
                     /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    1848..1965
                     /label=cPPT/CTS
                     /note="central polypurine tract and central termination 
                     sequence of HIV-1"
     promoter        2016..2264
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     2016..2036
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     2187..2206
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        2286..2361
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        2423..2634
                     /label=EF-1-alpha core promoter
                     /note="core promoter for human elongation factor 
                     EF-1-alpha"
     regulatory      2653..2662
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2659..6762
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             2659..6762
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             6763..6810
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             6811..6834
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             6844..6900
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             6901..7497
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="TEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIERV
                     TELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLAA
                     QQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETSA
                     PRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     7394..7414
                     /label=Puro-F
                     /note="Puromycin resistance gene, forward primer"
     misc_feature    7513..8101
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(7566..7586)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(7984..7995)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     LTR             8173..8406
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     primer_bind     complement(8432..8449)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    8438..8662
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      8708..9136
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(8795..8814)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     9005..9026
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(9145..9165)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        9150..9479
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      9330..9465
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     9392..9411
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        9527..9574
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             9593..9967
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     polyA_signal    10097..10218
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(10134..10153)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     10188..10207
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(10267..10283)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(10267..10283)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(10280..10302)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    10291..10307
                     /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(10315..10321,10322..10339,10340..10345))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    10360..10381
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(10498..10515)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(10669..11257)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(10749..10768)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(11428..12219,12220..12288))
                     /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     12051..12070
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(12289..12393)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(12468..12487)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        join(12659..13013,1..25)
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
ORIGIN
        1 cgtattagtc atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg
       61 atagcggttt gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt
      121 gttttggcac caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac
      181 gcaaatgggc ggtaggcgtg tacggtggga ggtctatata agcagcgcgt tttgcctgta
      241 ctgggtctct ctggttagac cagatctgag cctgggagct ctctggctaa ctagggaacc
      301 cactgcttaa gcctcaataa agcttgcctt gagtgcttca agtagtgtgt gcccgtctgt
      361 tgtgtgactc tggtaactag agatccctca gaccctttta gtcagtgtgg aaaatctcta
      421 gcagtggcgc ccgaacaggg acttgaaagc gaaagggaaa ccagaggagc tctctcgacg
      481 caggactcgg cttgctgaag cgcgcacggc aagaggcgag gggcggcgac tggtgagtac
      541 gccaaaaatt ttgactagcg gaggctagaa ggagagagat gggtgcgaga gcgtcagtat
      601 taagcggggg agaattagat cgcgatggga aaaaattcgg ttaaggccag ggggaaagaa
      661 aaaatataaa ttaaaacata tagtatgggc aagcagggag ctagaacgat tcgcagttaa
      721 tcctggcctg ttagaaacat cagaaggctg tagacaaata ctgggacagc tacaaccatc
      781 ccttcagaca ggatcagaag aacttagatc attatataat acagtagcaa ccctctattg
      841 tgtgcatcaa aggatagaga taaaagacac caaggaagct ttagacaaga tagaggaaga
      901 gcaaaacaaa agtaagacca ccgcacagca agcggccgct gatcttcaga cctggaggag
      961 gagatatgag ggacaattgg agaagtgaat tatataaata taaagtagta aaaattgaac
     1021 cattaggagt agcacccacc aaggcaaaga gaagagtggt gcagagagaa aaaagagcag
     1081 tgggaatagg agctttgttc cttgggttct tgggagcagc aggaagcact atgggcgcag
     1141 cgtcaatgac gctgacggta caggccagac aattattgtc tggtatagtg cagcagcaga
     1201 acaatttgct gagggctatt gaggcgcaac agcatctgtt gcaactcaca gtctggggca
     1261 tcaagcagct ccaggcaaga atcctggctg tggaaagata cctaaaggat caacagctcc
     1321 tggggatttg gggttgctct ggaaaactca tttgcaccac tgctgtgcct tggaatgcta
     1381 gttggagtaa taaatctctg gaacagattt ggaatcacac gacctggatg gagtgggaca
     1441 gagaaattaa caattacaca agcttaatac actccttaat tgaagaatcg caaaaccagc
     1501 aagaaaagaa tgaacaagaa ttattggaat tagataaatg ggcaagtttg tggaattggt
     1561 ttaacataac aaattggctg tggtatataa aattattcat aatgatagta ggaggcttgg
     1621 taggtttaag aatagttttt gctgtacttt ctatagtgaa tagagttagg cagggatatt
     1681 caccattatc gtttcagacc cacctcccaa ccccgagggg acccgacagg cccgaaggaa
     1741 tagaagaaga aggtggagag agagacagag acagatccat tcgattagtg aacggatcgg
     1801 cactgcgtgc gccaattctg cagacaaatg gcagtattca tccacaattt taaaagaaaa
     1861 ggggggattg gggggtacag tgcaggggaa agaatagtag acataatagc aacagacata
     1921 caaactaaag aattacaaaa acaaattaca aaaattcaaa attttcgggt ttattacagg
     1981 gacagcagag atccagtttg gttaattaag gtaccgaggg cctatttccc atgattcctt
     2041 catatttgca tatacgatac aaggctgtta gagagataat tagaattaat ttgactgtaa
     2101 acacaaagat attagtacaa aatacgtgac gtagaaagta ataatttctt gggtagtttg
     2161 cagttttaaa attatgtttt aaaatggact atcatatgct taccgtaact tgaaagtatt
     2221 tcgatttctt ggctttatat atcttgtgga aaggacgaaa caccgccgcc tagaagcatt
     2281 tgcgggtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta tcaacttgaa
     2341 aaagtggcac cgagtcggtg cttttttgaa ttcgctagct aggtcttgaa aggagtggga
     2401 attggctccg gtgcccgtca gtgggcagag cgcacatcgc ccacagtccc cgagaagttg
     2461 gggggagggg tcggcaattg atccggtgcc tagagaaggt ggcgcggggt aaactgggaa
     2521 agtgatgtcg tgtactggct ccgccttttt cccgagggtg ggggagaacc gtatataagt
     2581 gcagtagtcg ccgtgaacgt tctttttcgc aacgggtttg ccgccagaac acaggaccgg
     2641 ttctagagcg ctgccaccat ggacaagaag tacagcatcg gcctggacat cggcaccaac
     2701 tctgtgggct gggccgtgat caccgacgag tacaaggtgc ccagcaagaa attcaaggtg
     2761 ctgggcaaca ccgaccggca cagcatcaag aagaacctga tcggagccct gctgttcgac
     2821 agcggcgaaa cagccgaggc cacccggctg aagagaaccg ccagaagaag atacaccaga
     2881 cggaagaacc ggatctgcta tctgcaagag atcttcagca acgagatggc caaggtggac
     2941 gacagcttct tccacagact ggaagagtcc ttcctggtgg aagaggataa gaagcacgag
     3001 cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa gtaccccacc
     3061 atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct gcggctgatc
     3121 tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga gggcgacctg
     3181 aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac ctacaaccag
     3241 ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat cctgtctgcc
     3301 agactgagca agagcagacg gctggaaaat ctgatcgccc agctgcccgg cgagaagaag
     3361 aatggcctgt tcggaaacct gattgccctg agcctgggcc tgacccccaa cttcaagagc
     3421 aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta cgacgacgac
     3481 ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct ggccgccaag
     3541 aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga gatcaccaag
     3601 gcccccctga gcgcctctat gatcaagaga tacgacgagc accaccagga cctgaccctg
     3661 ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt cttcgaccag
     3721 agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga gttctacaag
     3781 ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt gaagctgaac
     3841 agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc ccaccagatc
     3901 cacctgggag agctgcacgc cattctgcgg cggcaggaag atttttaccc attcctgaag
     3961 gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta cgtgggccct
     4021 ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga aaccatcacc
     4081 ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt catcgagcgg
     4141 atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca cagcctgctg
     4201 tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac cgagggaatg
     4261 agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct gctgttcaag
     4321 accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa aatcgagtgc
     4381 ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct gggcacatac
     4441 cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga aaacgaggac
     4501 attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat gatcgaggaa
     4561 cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct gaagcggcgg
     4621 agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg ggacaagcag
     4681 tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag aaacttcatg
     4741 cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc
     4801 ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc cgccattaag
     4861 aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat gggccggcac
     4921 aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca gaagggacag
     4981 aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct gggcagccag
     5041 atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct gtacctgtac
     5101 tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc
     5161 gactacgatg tggaccatat cgtgcctcag agctttctga aggacgactc catcgacaac
     5221 aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag
     5281 gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct gattacccag
     5341 agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact ggataaggcc
     5401 ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt ggcacagatc
     5461 ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg ggaagtgaaa
     5521 gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca gttttacaaa
     5581 gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga
     5641 accgccctga tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag
     5701 gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc
     5761 aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac cctggccaac
     5821 ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg
     5881 gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca agtgaatatc
     5941 gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct gcccaagagg
     6001 aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta cggcggcttc
     6061 gacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc
     6121 aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag aagcagcttc
     6181 gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa aaaggacctg
     6241 atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa gagaatgctg
     6301 gcctctgccg gcgaactgca gaagggaaac gaactggccc tgccctccaa atatgtgaac
     6361 ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga taatgagcag
     6421 aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga gcagatcagc
     6481 gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct gtccgcctac
     6541 aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca cctgtttacc
     6601 ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat cgaccggaag
     6661 aggtacacca gcaccaaaga ggtgctggac gccaccctga tccaccagag catcaccggc
     6721 ctgtacgaga cacggatcga cctgtctcag ctgggaggcg acaagcgacc tgccgccaca
     6781 aagaaggctg gacaggctaa gaagaagaaa gattacaaag acgatgacga taagggatcc
     6841 ggcgcaacaa acttctctct gctgaaacaa gccggagatg tcgaagagaa tcctggaccg
     6901 accgagtaca agcccacggt gcgcctcgcc acccgcgacg acgtccccag ggccgtacgc
     6961 accctcgccg ccgcgttcgc cgactacccc gccacgcgcc acaccgtcga tccggaccgc
     7021 cacatcgagc gggtcaccga gctgcaagaa ctcttcctca cgcgcgtcgg gctcgacatc
     7081 ggcaaggtgt gggtcgcgga cgacggcgcc gcggtggcgg tctggaccac gccggagagc
     7141 gtcgaagcgg gggcggtgtt cgccgagatc ggcccgcgca tggccgagtt gagcggttcc
     7201 cggctggccg cgcagcaaca gatggaaggc ctcctggcgc cgcaccggcc caaggagccc
     7261 gcgtggttcc tggccaccgt cggagtctcg cccgaccacc agggcaaggg tctgggcagc
     7321 gccgtcgtgc tccccggagt ggaggcggcc gagcgcgccg gggtgcccgc cttcctggag
     7381 acctccgcgc cccgcaacct ccccttctac gagcggctcg gcttcaccgt caccgccgac
     7441 gtcgaggtgc ccgaaggacc gcgcacctgg tgcatgaccc gcaagcccgg tgcctgaacg
     7501 cgttaagtcg acaatcaacc tctggattac aaaatttgtg aaagattgac tggtattctt
     7561 aactatgttg ctccttttac gctatgtgga tacgctgctt taatgccttt gtatcatgct
     7621 attgcttccc gtatggcttt cattttctcc tccttgtata aatcctggtt gctgtctctt
     7681 tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac
     7741 gcaaccccca ctggttgggg cattgccacc acctgtcagc tcctttccgg gactttcgct
     7801 ttccccctcc ctattgccac ggcggaactc atcgccgcct gccttgcccg ctgctggaca
     7861 ggggctcggc tgttgggcac tgacaattcc gtggtgttgt cggggaaatc atcgtccttt
     7921 ccttggctgc tcgcctgtgt tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc
     7981 ccttcggccc tcaatccagc ggaccttcct tcccgcggcc tgctgccggc tctgcggcct
     8041 cttccgcgtc ttcgccttcg ccctcagacg agtcggatct ccctttgggc cgcctccccg
     8101 cgtcgacttt aagaccaatg acttacaagg cagctgtaga tcttagccac tttttaaaag
     8161 aaaagggggg actggaaggg ctaattcact cccaacgaag acaagatctg ctttttgctt
     8221 gtactgggtc tctctggtta gaccagatct gagcctggga gctctctggc taactaggga
     8281 acccactgct taagcctcaa taaagcttgc cttgagtgct tcaagtagtg tgtgcccgtc
     8341 tgttgtgtga ctctggtaac tagagatccc tcagaccctt ttagtcagtg tggaaaatct
     8401 ctagcagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     8461 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     8521 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     8581 gggggtgggg tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct
     8641 ggggatgcgg tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg
     8701 tatccccacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     8761 gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt
     8821 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     8881 cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt
     8941 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     9001 aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt
     9061 gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa
     9121 aaatttaacg cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag
     9181 gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg
     9241 gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag
     9301 caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc
     9361 attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg
     9421 cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa
     9481 agctcccggg agcttgtata tccattttcg gatctgatca gcacgtgttg acaattaatc
     9541 atcggcatag tatatcggca tagtataata cgacaaggtg aggaactaaa ccatggccaa
     9601 gttgaccagt gccgttccgg tgctcaccgc gcgcgacgtc gccggagcgg tcgagttctg
     9661 gaccgaccgg ctcgggttct cccgggactt cgtggaggac gacttcgccg gtgtggtccg
     9721 ggacgacgtg accctgttca tcagcgcggt ccaggaccag gtggtgccgg acaacaccct
     9781 ggcctgggtg tgggtgcgcg gcctggacga gctgtacgcc gagtggtcgg aggtcgtgtc
     9841 cacgaacttc cgggacgcct ccgggccggc catgaccgag atcggcgagc agccgtgggg
     9901 gcgggagttc gccctgcgcg acccggccgg caactgcgtg cacttcgtgg ccgaggagca
     9961 ggactgacac gtgctacgag atttcgattc caccgccgcc ttctatgaaa ggttgggctt
    10021 cggaatcgtt ttccgggacg ccggctggat gatcctccag cgcggggatc tcatgctgga
    10081 gttcttcgcc caccccaact tgtttattgc agcttataat ggttacaaat aaagcaatag
    10141 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
    10201 actcatcaat gtatcttatc atgtctgtat accgtcgacc tctagctaga gcttggcgta
    10261 atcatggtca tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat
    10321 acgagccgga agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt
    10381 aattgcgttg cgctcactgc ccgctttcca gtcgggaaac ctgtcgtgcc agctgcatta
    10441 atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt gggcgctctt ccgcttcctc
    10501 gctcactgac tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa
    10561 ggcggtaata cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa
    10621 aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct
    10681 ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac
    10741 aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc
    10801 gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc
    10861 tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg
    10921 tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga
    10981 gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta acaggattag
    11041 cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta
    11101 cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag
    11161 agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg
    11221 caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac
    11281 ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc
    11341 aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag
    11401 tatatatgag taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc
    11461 agcgatctgt ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac
    11521 gatacgggag ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc
    11581 accggctcca gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg
    11641 tcctgcaact ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag
    11701 tagttcgcca gttaatagtt tgcgcaacgt tgttgccatt gctacaggca tcgtggtgtc
    11761 acgctcgtcg tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac
    11821 atgatccccc atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag
    11881 aagtaagttg gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac
    11941 tgtcatgcca tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg
    12001 agaatagtgt atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc
    12061 gccacatagc agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact
    12121 ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg
    12181 atcttcagca tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa
    12241 tgccgcaaaa aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt
    12301 tcaatattat tgaagcattt atcagggtta ttgtctcatg agcggataca tatttgaatg
    12361 tatttagaaa aataaacaaa taggggttcc gcgcacattt ccccgaaaag tgccacctga
    12421 cgtcgacgga tcgggagatc tcccgatccc ctatggtgca ctctcagtac aatctgctct
    12481 gatgccgcat agttaagcca gtatctgctc cctgcttgtg tgttggaggt cgctgagtag
    12541 tgcgcgagca aaatttaagc tacaacaagg caaggcttga ccgacaattg catgaagaat
    12601 ctgcttaggg ttaggcgttt tgcgctgctt cgcgatgtac gggccagata tacgcgttga
    12661 cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt tcatagccca
    12721 tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg accgcccaac
    12781 gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc aatagggact
    12841 ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc agtacatcaa
    12901 gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg gcccgcctgg
    12961 cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat cta
//
