LOCUS       sequence-370988-        8435 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  A human codon-optimized SpCas9 nickase and chimeric g2 guide RNA 
            expression plasmid..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8435)
  AUTHORS   Javaid N, Choi S
  TITLE     An Alternate Approach to Generate Induced Pluripotent Stem Cells 
            with Precise CRISPR/Cas9 Tool.
  JOURNAL   Stem Cells Int. 2022 Sep 22;2022:4537335. doi: 10.1155/2022/4537335.
            eCollection 2022.
  PUBMED    36187228
REFERENCE   2  (bases 1 to 8435)
  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: "Stem Cells 
            Int."; date: "2022-09-22"; volume: "2022:4537335. doi"; pages: " 
            10.1155/2022/4537335. eCollection 2022"
COMMENT     Sequence Label: pX335-U6-Chimeric_BB-CBh-hSpCas9n(D10A)_g2_CASH-1
FEATURES             Location/Qualifiers
     source          1..8435
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..249
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     1..21
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     172..191
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        270..345
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        442..727
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        729..1006
                     /label=chicken beta-actin promoter
     intron          1007..1234
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     CDS             1255..1281
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1285..1305
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1315..1338,1339..1341,1342..5415)
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             1315..5415
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             5419..5439
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(5476..5493)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5482..5689
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5698..5838
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5698..5827
                     /label=AAV2 ITR
     rep_origin      5913..6368
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6000..6019)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6210..6231
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6385..6404)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6504..6526
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6564..6582)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6650..6754
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6755..6823,6824..7615)
                     /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(6973..6992)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7786..8374
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8275..8294
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gagggcctat ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag
       61 ataattggaa ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga
      121 aagtaataat ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat
      181 atgcttaccg taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga
      241 cgaaacaccg tcctaagacc accctgagcg ttttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tgttttagag
      361 ctagaaatag caagttaaaa taaggctagt ccgtttttag cgcgtgcgcc aattctgcag
      421 acaaatggct ctagaggtac ccgttacata acttacggta aatggcccgc ctggctgacc
      481 gcccaacgac ccccgcccat tgacgtcaat agtaacgcca atagggactt tccattgacg
      541 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
      601 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attgtgccca
      661 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
      721 taccatggtc gaggtgagcc ccacgttctg cttcactctc cccatctccc ccccctcccc
      781 acccccaatt ttgtatttat ttatttttta attattttgt gcagcgatgg gggcgggggg
      841 gggggggggg cgcgcgccrg gggggggggg ggggggsggg ggggggggsg gggsgrggcg
      901 gagaggtgcg gcggcagcca atcagagcgg cgcgctccga aagtttcctt ttatggcgag
      961 gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg cgggcgggag tcgctgcgcg
     1021 ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc cgcccgcccc ggctctgact
     1081 gaccgcgtta ctcccacagg tgagcgggcg ggacggccct tctcctccgg gctgtaatta
     1141 gctgagcaag aggtaagggt ttaagggatg gttggttggt ggggtattaa tgtttaatta
     1201 cctggagcac ctgcctgaaa tcactttttt tcaggttgga ccggtgccac catgtaccca
     1261 tacgatgttc cagattacgc ttcgccgaag aaaaagcgca aggtcgaagc gtccgacaag
     1321 aagtacagca tcggcctggc catcggcacc aactctgtgg gctgggccgt gatcaccgac
     1381 gagtacaagg tgcccagcaa gaaattcaag gtgctgggca acaccgaccg gcacagcatc
     1441 aagaagaacc tgatcggagc cctgctgttc gacagcggcg aaacagccga ggccacccgg
     1501 ctgaagagaa ccgccagaag aagatacacc agacggaaga accggatctg ctatctgcaa
     1561 gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccacag actggaagag
     1621 tccttcctgg tggaagagga taagaagcac gagcggcacc ccatcttcgg caacatcgtg
     1681 gacgaggtgg cctaccacga gaagtacccc accatctacc acctgagaaa gaaactggtg
     1741 gacagcaccg acaaggccga cctgcggctg atctatctgg ccctggccca catgatcaag
     1801 ttccggggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga cgtggacaag
     1861 ctgttcatcc agctggtgca gacctacaac cagctgttcg aggaaaaccc catcaacgcc
     1921 agcggcgtgg acgccaaggc catcctgtct gccagactga gcaagagcag acggctggaa
     1981 aatctgatcg cccagctgcc cggcgagaag aagaatggcc tgttcggcaa cctgattgcc
     2041 ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga ggatgccaaa
     2101 ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc ccagatcggc
     2161 gaccagtacg ccgacctgtt tctggccgcc aagaacctgt ccgacgccat cctgctgagc
     2221 gacatcctga gagtgaacac cgagatcacc aaggcccccc tgagcgcctc tatgatcaag
     2281 agatacgacg agcaccacca ggacctgacc ctgctgaaag ctctcgtgcg gcagcagctg
     2341 cctgagaagt acaaagagat tttcttcgac cagagcaaga acggctacgc cggctacatt
     2401 gacggcggag ccagccagga agagttctac aagttcatca agcccatcct ggaaaagatg
     2461 gacggcaccg aggaactgct cgtgaagctg aacagagagg acctgctgcg gaagcagcgg
     2521 accttcgaca acggcagcat cccccaccag atccacctgg gagagctgca cgccattctg
     2581 cggcggcagg aagattttta cccattcctg aaggacaacc gggaaaagat cgagaagatc
     2641 ctgaccttcc gcatccccta ctacgtgggc cctctggcca ggggaaacag cagattcgcc
     2701 tggatgacca gaaagagcga ggaaaccatc accccctgga acttcgagga agtggtggac
     2761 aagggcgctt ccgcccagag cttcatcgag cggatgacca acttcgataa gaacctgccc
     2821 aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt gtataacgag
     2881 ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc ccgccttcct gagcggcgag
     2941 cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc ggaaagtgac cgtgaagcag
     3001 ctgaaagagg actacttcaa gaaaatcgag tgcttcgact ccgtggaaat ctccggcgtg
     3061 gaagatcggt tcaacgcctc cctgggcaca taccacgatc tgctgaaaat tatcaaggac
     3121 aaggacttcc tggacaatga ggaaaacgag gacattctgg aagatatcgt gctgaccctg
     3181 acactgtttg aggacagaga gatgatcgag gaacggctga aaacctatgc ccacctgttc
     3241 gacgacaaag tgatgaagca gctgaagcgg cggagataca ccggctgggg caggctgagc
     3301 cggaagctga tcaacggcat ccgggacaag cagtccggca agacaatcct ggatttcctg
     3361 aagtccgacg gcttcgccaa cagaaacttc atgcagctga tccacgacga cagcctgacc
     3421 tttaaagagg acatccagaa agcccaggtg tccggccagg gcgatagcct gcacgagcac
     3481 attgccaatc tggccggcag ccccgccatt aagaagggca tcctgcagac agtgaaggtg
     3541 gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg agaacatcgt gatcgaaatg
     3601 gccagagaga accagaccac ccagaaggga cagaagaaca gccgcgagag aatgaagcgg
     3661 atcgaagagg gcatcaaaga gctgggcagc cagatcctga aagaacaccc cgtggaaaac
     3721 acccagctgc agaacgagaa gctgtacctg tactacctgc agaatgggcg ggatatgtac
     3781 gtggaccagg aactggacat caaccggctg tccgactacg atgtggacca tatcgtgcct
     3841 cagagctttc tgaaggacga ctccatcgac aacaaggtgc tgaccagaag cgacaagaac
     3901 cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga agaagatgaa gaactactgg
     3961 cggcagctgc tgaacgccaa gctgattacc cagagaaagt tcgacaatct gaccaaggcc
     4021 gagagaggcg gcctgagcga actggataag gccggcttca tcaagagaca gctggtggaa
     4081 acccggcaga tcacaaagca cgtggcacag atcctggact cccggatgaa cactaagtac
     4141 gacgagaatg acaagctgat ccgggaagtg aaagtgatca ccctgaagtc caagctggtg
     4201 tccgatttcc ggaaggattt ccagttttac aaagtgcgcg agatcaacaa ctaccaccac
     4261 gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc tgatcaaaaa gtaccctaag
     4321 ctggaaagcg agttcgtgta cggcgactac aaggtgtacg acgtgcggaa gatgatcgcc
     4381 aagagcgagc aggaaatcgg caaggctacc gccaagtact tcttctacag caacatcatg
     4441 aactttttca agaccgagat taccctggcc aacggcgaga tccggaagcg gcctctgatc
     4501 gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg gccgggattt tgccaccgtg
     4561 cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa agaccgaggt gcagacaggc
     4621 ggcttcagca aagagtctat cctgcccaag aggaacagcg ataagctgat cgccagaaag
     4681 aaggactggg accctaagaa gtacggcggc ttcgacagcc ccaccgtggc ctattctgtg
     4741 ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac tgaagagtgt gaaagagctg
     4801 ctggggatca ccatcatgga aagaagcagc ttcgagaaga atcccatcga ctttctggaa
     4861 gccaagggct acaaagaagt gaaaaaggac ctgatcatca agctgcctaa gtactccctg
     4921 ttcgagctgg aaaacggccg gaagagaatg ctggcctctg ccggcgaact gcagaaggga
     4981 aacgaactgg ccctgccctc caaatatgtg aacttcctgt acctggccag ccactatgag
     5041 aagctgaagg gctcccccga ggataatgag cagaaacagc tgtttgtgga acagcacaag
     5101 cactacctgg acgagatcat cgagcagatc agcgagttct ccaagagagt gatcctggcc
     5161 gacgctaatc tggacaaagt gctgtccgcc tacaacaagc accgggataa gcccatcaga
     5221 gagcaggccg agaatatcat ccacctgttt accctgacca atctgggagc ccctgccgcc
     5281 ttcaagtact ttgacaccac catcgaccgg aagaggtaca ccagcaccaa agaggtgctg
     5341 gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacacggat cgacctgtct
     5401 cagctgggag gcgacagccc caagaagaag agaaaggtgg aggccagcta agaattccta
     5461 gagctcgctg atcagcctcg actgtgcctt ctagttgcca gccatctgtt gtttgcccct
     5521 cccccgtgcc ttccttgacc ctggaaggtg ccactcccac tgtcctttcc taataaaatg
     5581 aggaaattgc atcgcattgt ctgagtaggt gtcattctat tctggggggt ggggtggggc
     5641 aggacagcaa gggggaggat tgggaagaga atagcaggca tgctggggag cggccgcagg
     5701 aacccctagt gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg
     5761 ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag
     5821 cgcgcagctg cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta
     5881 tttcacaccg catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc
     5941 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc
     6001 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     6061 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     6121 acttgatttg ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     6181 tttgacgttg gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact
     6241 caactctatc tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg
     6301 gttaaaaaat gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt
     6361 tacaatttta tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc
     6421 ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc
     6481 ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc
     6541 accgaaacgc gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
     6601 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
     6661 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     6721 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     6781 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     6841 gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct
     6901 caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac
     6961 ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact
     7021 cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa
     7081 gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga
     7141 taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt
     7201 tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga
     7261 agccatacca aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg
     7321 caaactatta actggcgaac tacttactct agcttcccgg caacaattaa tagactggat
     7381 ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat
     7441 tgctgataaa tctggagccg gtgagcgtgg aagccgcggt atcattgcag cactggggcc
     7501 agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga
     7561 tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc
     7621 agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag
     7681 gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc
     7741 gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt
     7801 tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt
     7861 gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat
     7921 accaaatact gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc
     7981 accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa
     8041 gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg
     8101 ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag
     8161 atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag
     8221 gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa
     8281 cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt
     8341 gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg
     8401 gttcctggcc ttttgctggc cttttgctca catgt
//
