LOCUS       sequence-252508-       10327 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  GFP positive gRNA/hSpCas9 construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10327)
  AUTHORS   Giacalone JC, Sharma TP, Burnight ER, Fingert JF, Mullins RF, Stone 
            EM, Tucker BA
  TITLE     CRISPR-Cas9-Based Genome Editing of Human Induced Pluripotent Stem 
            Cells.
  JOURNAL   Curr Protoc Stem Cell Biol. 2018 Feb 28;44:5B.7.1-5B.7.22. doi: 
            10.1002/cpsc.46.
  PUBMED    29512106
REFERENCE   2  (bases 1 to 10327)
  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: "Curr Protoc
            Stem Cell Biol."; date: "2018-02-28"; volume: "44:5B.7.1-5B.7.22. 
            doi"; pages: " 10.1002/cpsc.46"
COMMENT     Sequence Label: pX330GFP-hU6-gRNA-hSpCas9
FEATURES             Location/Qualifiers
     source          1..10327
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..286
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     intron          545..772
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      784..793
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             796..816
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             841..4941
                     /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             841..4941
                     /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             4942..4989
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5017..5034)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5023..5230
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5239..5379
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5239..5368
                     /label=AAV2 ITR
     rep_origin      5454..5909
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5541..5560)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5751..5772
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5926..5945)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6045..6067
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6105..6123)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6191..6295
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6296..6364,6365..7156)
                     /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(6514..6533)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7327..7915
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7816..7835
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        8114..8493
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        8494..8697
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     8648..8668
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     intron          8789..8921
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     CDS             8957..9622
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina 
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /label=CopGFP
                     /translation="MPAMKIECRITGTLNGVEFELVGGGEGTPEQGRMTNKMKSTKGAL
                     TFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYR
                     YEAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDG
                     GYYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA
                     "
     CDS             8957..9622
                     /codon_start=1
                     /product="green fluorescent protein 2 from Pontellina 
                     plumata, also known as ppluGFP2 (Shagin et al., 2004)"
                     /label=CopGFP
                     /translation="MPAMKIECRITGTLNGVEFELVGGGEGTPEQGRMTNKMKSTKGAL
                     TFSPYLLSHVMGYGFYHFGTYPSGYENPFLHAINNGGYTNTRIEKYEDGGVLHVSFSYR
                     YEAGRVIGDFKVVGTGFPEDSVIFTDKIIRSNATVEHLHPMGDNVLVGSFARTFSLRDG
                     GYYSFVVDSHMHFKSAIHPSILQNGGPMFAFRRVEELHSNTELGIVEYQHAFKTPIAFA
                     "
     primer_bind     complement(9636..9655)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     polyA_signal    9665..9746
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     9690..9709
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     promoter        9889..10137
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     9889..9909
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     10060..10079
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        10156..10231
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata gtaacgccaa tagggacttt ccattgacgt caatgggtgg agtatttacg
      121 gtaaactgcc cacttggcag tacatcaagt gtatcatatg ccaagtacgc cccctattga
      181 cgtcaatgac ggtaaatggc ccgcctggca ttgtgcccag tacatgacct tatgggactt
      241 tcctacttgg cagtacatct acgtattagt catcgctatt accatggtcg aggtgagccc
      301 cacgttctgc ttcactctcc ccatctcccc cccctcccca cccccaattt tgtatttatt
      361 tattttttaa ttattttgtg cagcgatggg ggcggggggg gggggggggg ggggcggggg
      421 gggggggggg gsgrggcggr grggkgcggc ggcagccaat cagagcggcg cgctccgaaa
      481 gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg
      541 ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg
      601 cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc
      661 tcctccgggc tgtaattagc tgagcaagag gtaagggttt aagggatggt tggttggtgg
      721 ggtattaatg tttaattacc tggagcacct gcctgaaatc actttttttc aggttggacc
      781 ggtgccacca tggccccaaa gaagaagcgg aaggtcggta tccacggagt cccagcagcc
      841 gacaagaagt acagcatcgg cctggacatc ggcaccaact ctgtgggctg ggccgtgatc
      901 accgacgagt acaaggtgcc cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac
      961 agcatcaaga agaacctgat cggagccctg ctgttcgaca gcggcgaaac agccgaggcc
     1021 acccggctga agagaaccgc cagaagaaga tacaccagac ggaagaaccg gatctgctat
     1081 ctgcaagaga tcttcagcaa cgagatggcc aaggtggacg acagcttctt ccacagactg
     1141 gaagagtcct tcctggtgga agaggataag aagcacgagc ggcaccccat cttcggcaac
     1201 atcgtggacg aggtggccta ccacgagaag taccccacca tctaccacct gagaaagaaa
     1261 ctggtggaca gcaccgacaa ggccgacctg cggctgatct atctggccct ggcccacatg
     1321 atcaagttcc ggggccactt cctgatcgag ggcgacctga accccgacaa cagcgacgtg
     1381 gacaagctgt tcatccagct ggtgcagacc tacaaccagc tgttcgagga aaaccccatc
     1441 aacgccagcg gcgtggacgc caaggccatc ctgtctgcca gactgagcaa gagcagacgg
     1501 ctggaaaatc tgatcgccca gctgcccggc gagaagaaga atggcctgtt cggaaacctg
     1561 attgccctga gcctgggcct gacccccaac ttcaagagca acttcgacct ggccgaggat
     1621 gccaaactgc agctgagcaa ggacacctac gacgacgacc tggacaacct gctggcccag
     1681 atcggcgacc agtacgccga cctgtttctg gccgccaaga acctgtccga cgccatcctg
     1741 ctgagcgaca tcctgagagt gaacaccgag atcaccaagg cccccctgag cgcctctatg
     1801 atcaagagat acgacgagca ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag
     1861 cagctgcctg agaagtacaa agagattttc ttcgaccaga gcaagaacgg ctacgccggc
     1921 tacattgacg gcggagccag ccaggaagag ttctacaagt tcatcaagcc catcctggaa
     1981 aagatggacg gcaccgagga actgctcgtg aagctgaaca gagaggacct gctgcggaag
     2041 cagcggacct tcgacaacgg cagcatcccc caccagatcc acctgggaga gctgcacgcc
     2101 attctgcggc ggcaggaaga tttttaccca ttcctgaagg acaaccggga aaagatcgag
     2161 aagatcctga ccttccgcat cccctactac gtgggccctc tggccagggg aaacagcaga
     2221 ttcgcctgga tgaccagaaa gagcgaggaa accatcaccc cctggaactt cgaggaagtg
     2281 gtggacaagg gcgcttccgc ccagagcttc atcgagcgga tgaccaactt cgataagaac
     2341 ctgcccaacg agaaggtgct gcccaagcac agcctgctgt acgagtactt caccgtgtat
     2401 aacgagctga ccaaagtgaa atacgtgacc gagggaatga gaaagcccgc cttcctgagc
     2461 ggcgagcaga aaaaggccat cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg
     2521 aagcagctga aagaggacta cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc
     2581 ggcgtggaag atcggttcaa cgcctccctg ggcacatacc acgatctgct gaaaattatc
     2641 aaggacaagg acttcctgga caatgaggaa aacgaggaca ttctggaaga tatcgtgctg
     2701 accctgacac tgtttgagga cagagagatg atcgaggaac ggctgaaaac ctatgcccac
     2761 ctgttcgacg acaaagtgat gaagcagctg aagcggcgga gatacaccgg ctggggcagg
     2821 ctgagccgga agctgatcaa cggcatccgg gacaagcagt ccggcaagac aatcctggat
     2881 ttcctgaagt ccgacggctt cgccaacaga aacttcatgc agctgatcca cgacgacagc
     2941 ctgaccttta aagaggacat ccagaaagcc caggtgtccg gccagggcga tagcctgcac
     3001 gagcacattg ccaatctggc cggcagcccc gccattaaga agggcatcct gcagacagtg
     3061 aaggtggtgg acgagctcgt gaaagtgatg ggccggcaca agcccgagaa catcgtgatc
     3121 gaaatggcca gagagaacca gaccacccag aagggacaga agaacagccg cgagagaatg
     3181 aagcggatcg aagagggcat caaagagctg ggcagccaga tcctgaaaga acaccccgtg
     3241 gaaaacaccc agctgcagaa cgagaagctg tacctgtact acctgcagaa tgggcgggat
     3301 atgtacgtgg accaggaact ggacatcaac cggctgtccg actacgatgt ggaccatatc
     3361 gtgcctcaga gctttctgaa ggacgactcc atcgacaaca aggtgctgac cagaagcgac
     3421 aagaaccggg gcaagagcga caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac
     3481 tactggcggc agctgctgaa cgccaagctg attacccaga gaaagttcga caatctgacc
     3541 aaggccgaga gaggcggcct gagcgaactg gataaggccg gcttcatcaa gagacagctg
     3601 gtggaaaccc ggcagatcac aaagcacgtg gcacagatcc tggactcccg gatgaacact
     3661 aagtacgacg agaatgacaa gctgatccgg gaagtgaaag tgatcaccct gaagtccaag
     3721 ctggtgtccg atttccggaa ggatttccag ttttacaaag tgcgcgagat caacaactac
     3781 caccacgccc acgacgccta cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac
     3841 cctaagctgg aaagcgagtt cgtgtacggc gactacaagg tgtacgacgt gcggaagatg
     3901 atcgccaaga gcgagcagga aatcggcaag gctaccgcca agtacttctt ctacagcaac
     3961 atcatgaact ttttcaagac cgagattacc ctggccaacg gcgagatccg gaagcggcct
     4021 ctgatcgaga caaacggcga aaccggggag atcgtgtggg ataagggccg ggattttgcc
     4081 accgtgcgga aagtgctgag catgccccaa gtgaatatcg tgaaaaagac cgaggtgcag
     4141 acaggcggct tcagcaaaga gtctatcctg cccaagagga acagcgataa gctgatcgcc
     4201 agaaagaagg actgggaccc taagaagtac ggcggcttcg acagccccac cgtggcctat
     4261 tctgtgctgg tggtggccaa agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa
     4321 gagctgctgg ggatcaccat catggaaaga agcagcttcg agaagaatcc catcgacttt
     4381 ctggaagcca agggctacaa agaagtgaaa aaggacctga tcatcaagct gcctaagtac
     4441 tccctgttcg agctggaaaa cggccggaag agaatgctgg cctctgccgg cgaactgcag
     4501 aagggaaacg aactggccct gccctccaaa tatgtgaact tcctgtacct ggccagccac
     4561 tatgagaagc tgaagggctc ccccgaggat aatgagcaga aacagctgtt tgtggaacag
     4621 cacaagcact acctggacga gatcatcgag cagatcagcg agttctccaa gagagtgatc
     4681 ctggccgacg ctaatctgga caaagtgctg tccgcctaca acaagcaccg ggataagccc
     4741 atcagagagc aggccgagaa tatcatccac ctgtttaccc tgaccaatct gggagcccct
     4801 gccgccttca agtactttga caccaccatc gaccggaaga ggtacaccag caccaaagag
     4861 gtgctggacg ccaccctgat ccaccagagc atcaccggcc tgtacgagac acggatcgac
     4921 ctgtctcagc tgggaggcga caaaaggccg gcggccacga aaaaggccgg ccaggcaaaa
     4981 aagaaaaagt aagaattcct agagctcgct gatcagcctc gactgtgcct tctagttgcc
     5041 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     5101 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     5161 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagag aatagcaggc
     5221 atgctgggga gcggccgcag gaacccctag tgatggagtt ggccactccc tctctgcgcg
     5281 ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc tttgcccggg
     5341 cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg cggtattttc
     5401 tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat agtacgcgcc
     5461 ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact
     5521 tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc
     5581 cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt
     5641 acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc
     5701 ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt
     5761 gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt tataagggat
     5821 tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa
     5881 ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa tctgctctga
     5941 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     6001 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     6061 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     6121 atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg gcacttttcg
     6181 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     6241 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     6301 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     6361 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     6421 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     6481 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     6541 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     6601 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     6661 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     6721 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     6781 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     6841 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     6901 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     6961 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg gaagccgcgg
     7021 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     7081 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     7141 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     7201 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     7261 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     7321 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     7381 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     7441 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     7501 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     7561 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     7621 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     7681 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     7741 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     7801 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     7861 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     7921 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttcaa
     7981 tattggccat tagccatatt attcattggt tatatagcat aaatcaatat tggctattgg
     8041 ccattgcata cgttgtatct atatcataat atgtacattt atattggctc atgtccaata
     8101 tgaccgccat gttggcattg attattgact agttattaat agtaatcaat tacggggtca
     8161 ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct
     8221 ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta
     8281 acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta aactgcccac
     8341 ttggcagtac atcaagtgta tcatatgcca agtccgcccc ctattgacgt caatgacggt
     8401 aaatggcccg cctggcatta tgcccagtac atgaccttac gggactttcc tacttggcag
     8461 tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca gtacaccaat
     8521 gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat tgacgtcaat
     8581 gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa taaccccgcc
     8641 ccgttgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag cagaggtcgt
     8701 ttagtgaacc gtcagatcac tagtagcttt attgcggtag tttatcacag ttaaattgct
     8761 aacgcagtca gtgctcgact gatcacaggt aagtatcaag gttacaagac aggtttaagg
     8821 aggccaatag aaactgggct tgtcgagaca gagaagattc ttgcgtttct gataggcacc
     8881 tattggtctt actgacatcc actttgcctt tctctccaca ggggtaccga agccgctagc
     8941 gctaccggtc gccaccatgc ccgccatgaa gatcgagtgc cgcatcaccg gcaccctgaa
     9001 cggcgtggag ttcgagctgg tgggcggcgg agagggcacc cccgagcagg gccgcatgac
     9061 caacaagatg aagagcacca aaggcgccct gaccttcagc ccctacctgc tgagccacgt
     9121 gatgggctac ggcttctacc acttcggcac ctaccccagc ggctacgaga accccttcct
     9181 gcacgccatc aacaacggcg gctacaccaa cacccgcatc gagaagtacg aggacggcgg
     9241 cgtgctgcac gtgagcttca gctaccgcta cgaggccggc cgcgtgatcg gcgacttcaa
     9301 ggtggtgggc accggcttcc ccgaggacag cgtgatcttc accgacaaga tcatccgcag
     9361 caacgccacc gtggagcacc tgcaccccat gggcgataac gtgctggtgg gcagcttcgc
     9421 ccgcaccttc agcctgcgcg acggcggcta ctacagcttc gtggtggaca gccacatgca
     9481 cttcaagagc gccatccacc ccagcatcct gcagaacggg ggccccatgt tcgccttccg
     9541 ccgcgtggag gagctgcaca gcaacaccga gctgggcatc gtggagtacc agcacgcctt
     9601 caagaccccc atcgccttcg ccagatctcg agctcgatga gtttggacaa accacaacta
     9661 gaatgcagtg aaaaaaatgc tttatttgtg aaatttgtga tgctattgct ttatttgtaa
     9721 ccattataag ctgcaataaa caagttaaca acaacaattg cattcatttt atgtttcagg
     9781 ttcaggggga ggtgtgggag gttttttaaa gcaagtaaaa cctctacaaa tgtggtactt
     9841 aagaggggga gaccaaaggg cgagacgtta aggcctcacg tgacatgtga gggcctattt
     9901 cccatgattc cttcatattt gcatatacga tacaaggctg ttagagagat aattggaatt
     9961 aatttgactg taaacacaaa gatattagta caaaatacgt gacgtagaaa gtaataattt
    10021 cttgggtagt ttgcagtttt aaaattatgt tttaaaatgg actatcatat gcttaccgta
    10081 acttgaaagt atttcgattt cttggcttta tatatcttgt ggaaaggacg aaacaccggg
    10141 tcttcgagaa gacctgtttt agagctagaa atagcaagtt aaaataaggc tagtccgtta
    10201 tcaacttgaa aaagtggcac cgagtcggtg cttttttgtt ttagagctag aaatagcaag
    10261 ttaaaataag gctagtccgt ttttagcgcg tgcgccaatt ctgcagacaa atggctctag
    10321 aggtacc
//
