LOCUS       sequence-106037-       10203 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  This plasmid separately encodes a human codon-optimized SpCas9, a 
            tracrRNA and customizable crRNA..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10203)
  AUTHORS   Cong L, Ran FA, Cox D, Lin S, Barretto R, Habib N, Hsu PD, Wu X, 
            Jiang W, Marraffini LA, Zhang F
  TITLE     Multiplex Genome Engineering Using CRISPR/Cas Systems.
  JOURNAL   Science. 2013 Jan 3.
  PUBMED    23287718
REFERENCE   2  (bases 1 to 10203)
  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: "Science. 
            2013 Jan 3."
COMMENT     Sequence Label: 
            pX260-U6-DR-BB-DR-Cbh-NLS-hSpCas9-NLS-H1-shorttracr-PGK-puro
FEATURES             Location/Qualifiers
     source          1..10203
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        157..442
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     repeat_region   493..528
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     repeat_region   545..580
                     /label=DR
                     /note="direct repeat for the Streptococcus pyogenes 
                     CRISPR/Cas system"
     promoter        complement(583..831)
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     complement(641..660)
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     primer_bind     complement(811..831)
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     rep_origin      complement(893..1481)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(973..992)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1652..2443,2444..2512))
                     /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     2275..2294
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2513..2617)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     2685..2703
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(2741..2763)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     2863..2882
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(2899..3354)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3036..3057)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3248..3267
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     CDS             complement(3627..4226)
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APHNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     primer_bind     4207..4226
                     /label=Puro-R
                     /note="Puromycin resistance gene, reverse primer. Also 
                     called puro-variant-R"
     promoter        complement(4266..4765)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     primer_bind     complement(4314..4333)
                     /label=mPGK-F
                     /note="Mouse PGK promoter, forward primer"
     primer_bind     4707..4730
                     /label=MSCV-rev
                     /note="Murine stem cell virus, reverse primer"
     protein_bind    complement(4826..4859)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(4935..4956)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_RNA        complement(5030..5108)
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     promoter        complement(5119..5333)
                     /label=H1 promoter
                     /note="human H1 RNA promoter"
     primer_bind     complement(5179..5198)
                     /label=H1
                     /note="Human H1 promoter, forward primer"
     polyA_signal    5347..5554
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     5543..5560
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(5588..5635)
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             complement(5636..9736)
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(5636..9736)
                     /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="DKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             complement(9761..9781)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(9788..9853)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     regulatory      9853..9862
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     intron          9874..10101
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     promoter        complement(join(10102..10203,1..155))
                     /label=chicken beta-actin promoter
                     /note="contains a 21-bp deletion relative to the standard 
                     chicken beta-actin promoter"
ORIGIN
        1 tctccgcctc gccccgcccc gcccctcgcc ccgccccccc cccccccccg cccccatcgc
       61 tgcacaaaat aattaaaaaa taaataaata caaaattggg ggtggggagg ggggggagat
      121 ggggagagtg aagcagaacg tggggctcac ctcgaccatg gtaatagcga tgactaatac
      181 gtagatgtac tgccaagtag gaaagtccca taaggtcatg tactgggcac aatgccaggc
      241 gggccattta ccgtcattga cgtcaatagg gggcgtactt ggcatatgat acacttgatg
      301 tactgccaag tgggcagttt accgtaaata ctccacccat tgacgtcaat ggaaagtccc
      361 tattggcgtt actattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc
      421 atttaccgta agttatgtaa cgggtacctc tagagccatt tgtctgcaga attggcgcac
      481 gcgctagaaa aagttttggg accattcaaa acagcatagc tctaaaacgt cttctcgaag
      541 acccgttttg ggaccattca aaacagcata gctctaaaac ccggtgtttc gtcctttcca
      601 caagatatat aaagccaaga aatcgaaata ctttcaagtt acggtaagca tatgatagtc
      661 cattttaaaa cataatttta aaactgcaaa ctacccaaga aattattact ttctacgtca
      721 cgtattttgt actaatatct ttgtgtttac agtcaaatta attccaatta tctctctaac
      781 agccttgtat cgtatatgca aatatgaagg aatcatggga aataggccct cacatgtgag
      841 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
      901 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
      961 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     1021 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     1081 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     1141 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     1201 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     1261 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     1321 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     1381 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     1441 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     1501 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     1561 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     1621 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     1681 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     1741 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cggcttccac
     1801 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     1861 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     1921 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     1981 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     2041 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     2101 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     2161 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     2221 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     2281 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     2341 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     2401 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     2461 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     2521 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     2581 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     2641 ctgacgtcta agaaaccatt attatcatga cattaaccta taaaaatagg cgtatcacga
     2701 ggccctttcg tctcgcgcgt ttcggtgatg acggtgaaaa cctctgacac atgcagctcc
     2761 cggagacggt cacagcttgt ctgtaagcgg atgccgggag cagacaagcc cgtcagggcg
     2821 cgtcagcggg tgttggcggg tgtcggggct ggcttaacta tgcggcatca gagcagattg
     2881 tactgagagt gcaccataaa attgtaaacg ttaatatttt gttaaaattc gcgttaaatt
     2941 tttgttaaat cagctcattt tttaaccaat agaccgaaat cggcaaaatc ccttataaat
     3001 caaaagaata gcccgagata gagttgagtg ttgttccagt ttggaacaag agtccactat
     3061 taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc gatggcccac
     3121 tacgtgaacc atcacccaaa tcaagttttt tggggtcgag gtgccgtaaa gcactaaatc
     3181 ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg aacgtggcga
     3241 gaaaggaagg gaagaaagcg aaaggagcgg gcgctaaggc gctggcaagt gtagcggtca
     3301 cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc gcgtactatg
     3361 gttgctttga cgtatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat
     3421 caggcgccgc tccaatcctt ccattcaaaa tgtaggtact ctgttctcac ccttcttaac
     3481 aaagtatgac aggaaaaact tccattttag tggacatctt tattgtttaa tagatcatca
     3541 atttctgcat cccggggatc tgatatcatc gatgcatggg gtcgtgcgct cctttcggtc
     3601 gggcgctgcg ggtcgtgggg cgggcgtcag gcaccgggct tgcgggtcat gcaccaggtg
     3661 cgcggtcctt cgggcacctc gacgtcggcg gtgacggtga agccgagccg ctcgtagaag
     3721 gggaggttgt ggggcgcgga ggtctccagg aaggcgggca ccccggcgcg ctcggccgcc
     3781 tccactccgg ggagcacgac ggcgctgccc agacccttgc cctggtggtc gggcgagacg
     3841 ccgacggtgg ccaggaacca cgcgggctcc ttgggccggt gcggcgccag gaggccttcc
     3901 atctgttgct gcgcggccag ccgggaaccg ctcaactcgg ccatgcgcgg gccgatctcg
     3961 gcgaacaccg cccccgcttc gacgctctcc ggcgtggtcc agaccgccac cgcggcgccg
     4021 tcgtccgcga cccacacctt gccgatgtcg agcccgacgc gcgtgaggaa gagttcttgc
     4081 agctcggtga cccgctcgat gtggcggtcc ggatcgacgg tgtggcgcgt ggcggggtag
     4141 tcggcgaacg cggcggcgag ggtgcgtacg gccctgggga cgtcgtcgcg ggtggcgagg
     4201 cgcaccgtgg gcttgtactc ggtcatggta agcttcagct gctcgagatc tagatggatg
     4261 caggtcgaaa ggcccggaga tgaggaagag gagaacagcg cggcagacgt gcgcttttga
     4321 agcgtgcaga atgccgggcc tccggaggac cttcgggcgc ccgccccgcc cctgagcccg
     4381 cccctgagcc cgcccccgga cccacccctt cccagcctct gagcccagaa agcgaaggag
     4441 caaagctgct attggccgct gccccaaagg cctacccgct tccattgctc agcggtgctg
     4501 tccatctgca cgagactagt gagacgtgct acttccattt gtcacgtcct gcacgacgcg
     4561 agctgcgggg cgggggggaa cttcctgact aggggaggag tagaaggtgg cgcgaagggg
     4621 ccaccaaaga acggagccgg ttggcgccta ccggtggatg tggaatgtgt gcgaggccag
     4681 aggccacttg tgtagcgcca agtgcccagc ggggctgcta aagcgcatgc tccagactgc
     4741 cttgggaaaa gcgcctcccc tacccggtag aatttcgagg tcgagatcct aagcttggct
     4801 ggacgtaaac tcctcttcag acctaataac ttcgtatagc atacattata cgaagttata
     4861 ttaagggtta ttgaatatga tcggaattct tacttgtaca gctcgtccat gccgagagtg
     4921 atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc gttggggtct
     4981 ttgctcaggg cggactgggt gctcaggtag tggttgtcgg cgccaaaaag caccgactcg
     5041 gtgccacttt ttcaagttga taacggacta gccttatttt aacttgctat gctgttttga
     5101 atggttccaa ctagatctgt ggtctcatac agaacttata agattcccaa atccaaagac
     5161 atttcacgtt tatggtgatt tcccagaaca catagcgaca tgcaaatatt gcagggcgcc
     5221 actcccctgt ccctcacagc catcttcctg ccagggcgca cgcgcgctgg gtgttcccgc
     5281 ctagtgacac tgggcccgcg attccttgga gcgggttgat gacgtcagcg ttcgaattgc
     5341 ggccgctccc cagcatgcct gctattctct tcccaatcct cccccttgct gtcctgcccc
     5401 accccacccc ccagaataga atgacaccta ctcagacaat gcgatgcaat ttcctcattt
     5461 tattaggaaa ggacagtggg agtggcacct tccagggtca aggaaggcac gggggagggg
     5521 caaacaacag atggctggca actagaaggc acagtcgagg ctgatcagcg agctctagga
     5581 attcttactt tttctttttt gcctggccgg cctttttcgt ggccgccggc cttttgtcgc
     5641 ctcccagctg agacaggtcg atccgtgtct cgtacaggcc ggtgatgctc tggtggatca
     5701 gggtggcgtc cagcacctct ttggtgctgg tgtacctctt ccggtcgatg gtggtgtcaa
     5761 agtacttgaa ggcggcaggg gctcccagat tggtcagggt aaacaggtgg atgatattct
     5821 cggcctgctc tctgatgggc ttatcccggt gcttgttgta ggcggacagc actttgtcca
     5881 gattagcgtc ggccaggatc actctcttgg agaactcgct gatctgctcg atgatctcgt
     5941 ccaggtagtg cttgtgctgt tccacaaaca gctgtttctg ctcattatcc tcgggggagc
     6001 ccttcagctt ctcatagtgg ctggccaggt acaggaagtt cacatatttg gagggcaggg
     6061 ccagttcgtt tcccttctgc agttcgccgg cagaggccag cattctcttc cggccgtttt
     6121 ccagctcgaa cagggagtac ttaggcagct tgatgatcag gtcctttttc acttctttgt
     6181 agcccttggc ttccagaaag tcgatgggat tcttctcgaa gctgcttctt tccatgatgg
     6241 tgatccccag cagctctttc acactcttca gtttcttgga cttgcccttt tccactttgg
     6301 ccaccaccag cacagaatag gccacggtgg ggctgtcgaa gccgccgtac ttcttagggt
     6361 cccagtcctt ctttctggcg atcagcttat cgctgttcct cttgggcagg atagactctt
     6421 tgctgaagcc gcctgtctgc acctcggtct ttttcacgat attcacttgg ggcatgctca
     6481 gcactttccg cacggtggca aaatcccggc ccttatccca cacgatctcc ccggtttcgc
     6541 cgtttgtctc gatcagaggc cgcttccgga tctcgccgtt ggccagggta atctcggtct
     6601 tgaaaaagtt catgatgttg ctgtagaaga agtacttggc ggtagccttg ccgatttcct
     6661 gctcgctctt ggcgatcatc ttccgcacgt cgtacacctt gtagtcgccg tacacgaact
     6721 cgctttccag cttagggtac tttttgatca gggcggttcc cacgacggcg ttcaggtagg
     6781 cgtcgtgggc gtggtggtag ttgttgatct cgcgcacttt gtaaaactgg aaatccttcc
     6841 ggaaatcgga caccagcttg gacttcaggg tgatcacttt cacttcccgg atcagcttgt
     6901 cattctcgtc gtacttagtg ttcatccggg agtccaggat ctgtgccacg tgctttgtga
     6961 tctgccgggt ttccaccagc tgtctcttga tgaagccggc cttatccagt tcgctcaggc
     7021 cgcctctctc ggccttggtc agattgtcga actttctctg ggtaatcagc ttggcgttca
     7081 gcagctgccg ccagtagttc ttcatcttct tcacgacctc ttcggagggc acgttgtcgc
     7141 tcttgccccg gttcttgtcg cttctggtca gcaccttgtt gtcgatggag tcgtccttca
     7201 gaaagctctg aggcacgata tggtccacat cgtagtcgga cagccggttg atgtccagtt
     7261 cctggtccac gtacatatcc cgcccattct gcaggtagta caggtacagc ttctcgttct
     7321 gcagctgggt gttttccacg gggtgttctt tcaggatctg gctgcccagc tctttgatgc
     7381 cctcttcgat ccgcttcatt ctctcgcggc tgttcttctg tcccttctgg gtggtctggt
     7441 tctctctggc catttcgatc acgatgttct cgggcttgtg ccggcccatc actttcacga
     7501 gctcgtccac caccttcact gtctgcagga tgcccttctt aatggcgggg ctgccggcca
     7561 gattggcaat gtgctcgtgc aggctatcgc cctggccgga cacctgggct ttctggatgt
     7621 cctctttaaa ggtcaggctg tcgtcgtgga tcagctgcat gaagtttctg ttggcgaagc
     7681 cgtcggactt caggaaatcc aggattgtct tgccggactg cttgtcccgg atgccgttga
     7741 tcagcttccg gctcagcctg ccccagccgg tgtatctccg ccgcttcagc tgcttcatca
     7801 ctttgtcgtc gaacaggtgg gcataggttt tcagccgttc ctcgatcatc tctctgtcct
     7861 caaacagtgt cagggtcagc acgatatctt ccagaatgtc ctcgttttcc tcattgtcca
     7921 ggaagtcctt gtccttgata attttcagca gatcgtggta tgtgcccagg gaggcgttga
     7981 accgatcttc cacgccggag atttccacgg agtcgaagca ctcgattttc ttgaagtagt
     8041 cctctttcag ctgcttcacg gtcactttcc ggttggtctt gaacagcagg tccacgatgg
     8101 cctttttctg ctcgccgctc aggaaggcgg gctttctcat tccctcggtc acgtatttca
     8161 ctttggtcag ctcgttatac acggtgaagt actcgtacag caggctgtgc ttgggcagca
     8221 ccttctcgtt gggcaggttc ttatcgaagt tggtcatccg ctcgatgaag ctctgggcgg
     8281 aagcgccctt gtccaccact tcctcgaagt tccagggggt gatggtttcc tcgctctttc
     8341 tggtcatcca ggcgaatctg ctgtttcccc tggccagagg gcccacgtag taggggatgc
     8401 ggaaggtcag gatcttctcg atcttttccc ggttgtcctt caggaatggg taaaaatctt
     8461 cctgccgccg cagaatggcg tgcagctctc ccaggtggat ctggtggggg atgctgccgt
     8521 tgtcgaaggt ccgctgcttc cgcagcaggt cctctctgtt cagcttcacg agcagttcct
     8581 cggtgccgtc catcttttcc aggatgggct tgatgaactt gtagaactct tcctggctgg
     8641 ctccgccgtc aatgtagccg gcgtagccgt tcttgctctg gtcgaagaaa atctctttgt
     8701 acttctcagg cagctgctgc cgcacgagag ctttcagcag ggtcaggtcc tggtggtgct
     8761 cgtcgtatct cttgatcata gaggcgctca ggggggcctt ggtgatctcg gtgttcactc
     8821 tcaggatgtc gctcagcagg atggcgtcgg acaggttctt ggcggccaga aacaggtcgg
     8881 cgtactggtc gccgatctgg gccagcaggt tgtccaggtc gtcgtcgtag gtgtccttgc
     8941 tcagctgcag tttggcatcc tcggccaggt cgaagttgct cttgaagttg ggggtcaggc
     9001 ccaggctcag ggcaatcagg tttccgaaca ggccattctt cttctcgccg ggcagctggg
     9061 cgatcagatt ttccagccgt ctgctcttgc tcagtctggc agacaggatg gccttggcgt
     9121 ccacgccgct ggcgttgatg gggttttcct cgaacagctg gttgtaggtc tgcaccagct
     9181 ggatgaacag cttgtccacg tcgctgttgt cggggttcag gtcgccctcg atcaggaagt
     9241 ggccccggaa cttgatcatg tgggccaggg ccagatagat cagccgcagg tcggccttgt
     9301 cggtgctgtc caccagtttc tttctcaggt ggtagatggt ggggtacttc tcgtggtagg
     9361 ccacctcgtc cacgatgttg ccgaagatgg ggtgccgctc gtgcttctta tcctcttcca
     9421 ccaggaagga ctcttccagt ctgtggaaga agctgtcgtc caccttggcc atctcgttgc
     9481 tgaagatctc ttgcagatag cagatccggt tcttccgtct ggtgtatctt cttctggcgg
     9541 ttctcttcag ccgggtggcc tcggctgttt cgccgctgtc gaacagcagg gctccgatca
     9601 ggttcttctt gatgctgtgc cggtcggtgt tgcccagcac cttgaatttc ttgctgggca
     9661 ccttgtactc gtcggtgatc acggcccagc ccacagagtt ggtgccgatg tccaggccga
     9721 tgctgtactt cttgtcggct gctgggactc cgtggatacc gaccttccgc ttcttctttg
     9781 gggccatctt atcgtcatcg tctttgtaat caatatcatg atccttgtag tctccgtcgt
     9841 ggtccttata gtccatggtg gcaccggtcc aacctgaaaa aaagtgattt caggcaggtg
     9901 ctccaggtaa ttaaacatta ataccccacc aaccaaccat cccttaaacc cttacctctt
     9961 gctcagctaa ttacagcccg gaggagaagg gccgtcccgc ccgctcacct gtgggagtaa
    10021 cgcggtcagt cagagccggg gcgggcggcg cgaggcggcg gcggagcggg gcacggggcg
    10081 aaggcagcgc gcagcgactc ccgcccgccg cgcgcttcgc tttttatagg gccgccgccg
    10141 ccgccgcctc gccataaaag gaaactttcg gagcgcgccg ctctgattgg ctgccgccgc
    10201 acc
//
