LOCUS       sequence-242254-        8487 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  gRNA/Cas9 expression for 2nd editing using pSLHDR01, 02.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8487)
  AUTHORS   Ikeda K, Uchida N, Nishimura T, White J, Martin RM, Nakauchi H, 
            Sebastiano V, Weinberg KI, Porteus MH
  TITLE     Efficient scarless genome editing in human pluripotent stem cells.
  JOURNAL   Nat Methods. 2018 Dec;15(12):1045-1047. doi: 
            10.1038/s41592-018-0212-y. Epub 2018 Nov 30.
  PUBMED    30504872
REFERENCE   2  (bases 1 to 8487)
  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"; doi: 
            "10.1038/s41592-018-0212-y"; journalName: "Nat Methods"; date: 
            "2018-12"; volume: "15"; issue: "12"; pages: "1045-1047"
COMMENT     Sequence Label: pSLHDR_R1
FEATURES             Location/Qualifiers
     source          1..8487
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     intron          187..414
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      426..435
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             435..500
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             507..527
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             552..4652
                     /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             552..4652
                     /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             4653..4700
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(4728..4745)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4734..4941
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4950..5090
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4950..5079
                     /label=AAV2 ITR
     rep_origin      5165..5620
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5252..5271)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5462..5483
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5637..5656)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5756..5778
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5816..5834)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5902..6006
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6007..6075,6076..6867)
                     /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(6225..6244)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7038..7626
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7527..7546
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        7688..7936
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7688..7708
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7859..7878
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        7958..8033
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        8130..8415
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        join(8417..8487,1..186)
                     /label=chicken beta-actin promoter
                     /note="contains a 21-bp deletion relative to the standard 
                     chicken beta-actin promoter"
ORIGIN
        1 tttatttttt aattattttg tgcagcgatg ggggcggggg gggggggggg gcggggcgag
       61 ggggggggcg gggcggggcg gagaggtgcg gcggcagcca atcagagcgg cgcgctccga
      121 aagtttcctt ttatggcgag gcggcggcgg cggcggccct ataaaaagcg aagcgcgcgg
      181 cgggcgggag tcgctgcgcg ctgccttcgc cccgtgcccc gctccgccgc cgcctcgcgc
      241 cgcccgcccc ggctctgact gaccgcgtta ctcccacagg tgagcgggcg ggacggccct
      301 tctcctccgg gctgtaatta gctgagcaag aggtaagggt ttaagggatg gttggttggt
      361 ggggtattaa tgtttaatta cctggagcac ctgcctgaaa tcactttttt tcaggttgga
      421 ccggtgccac catggactat aaggaccacg acggagacta caaggatcat gatattgatt
      481 acaaagacga tgacgataag atggccccaa agaagaagcg gaaggtcggt atccacggag
      541 tcccagcagc cgacaagaag tacagcatcg gcctggacat cggcaccaac tctgtgggct
      601 gggccgtgat caccgacgag tacaaggtgc ccagcaagaa attcaaggtg ctgggcaaca
      661 ccgaccggca cagcatcaag aagaacctga tcggagccct gctgttcgac agcggcgaaa
      721 cagccgaggc cacccggctg aagagaaccg ccagaagaag atacaccaga cggaagaacc
      781 ggatctgcta tctgcaagag atcttcagca acgagatggc caaggtggac gacagcttct
      841 tccacagact ggaagagtcc ttcctggtgg aagaggataa gaagcacgag cggcacccca
      901 tcttcggcaa catcgtggac gaggtggcct accacgagaa gtaccccacc atctaccacc
      961 tgagaaagaa actggtggac agcaccgaca aggccgacct gcggctgatc tatctggccc
     1021 tggcccacat gatcaagttc cggggccact tcctgatcga gggcgacctg aaccccgaca
     1081 acagcgacgt ggacaagctg ttcatccagc tggtgcagac ctacaaccag ctgttcgagg
     1141 aaaaccccat caacgccagc ggcgtggacg ccaaggccat cctgtctgcc agactgagca
     1201 agagcagacg gctggaaaat ctgatcgccc agctgcccgg cgagaagaag aatggcctgt
     1261 tcggaaacct gattgccctg agcctgggcc tgacccccaa cttcaagagc aacttcgacc
     1321 tggccgagga tgccaaactg cagctgagca aggacaccta cgacgacgac ctggacaacc
     1381 tgctggccca gatcggcgac cagtacgccg acctgtttct ggccgccaag aacctgtccg
     1441 acgccatcct gctgagcgac atcctgagag tgaacaccga gatcaccaag gcccccctga
     1501 gcgcctctat gatcaagaga tacgacgagc accaccagga cctgaccctg ctgaaagctc
     1561 tcgtgcggca gcagctgcct gagaagtaca aagagatttt cttcgaccag agcaagaacg
     1621 gctacgccgg ctacattgac ggcggagcca gccaggaaga gttctacaag ttcatcaagc
     1681 ccatcctgga aaagatggac ggcaccgagg aactgctcgt gaagctgaac agagaggacc
     1741 tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc ccaccagatc cacctgggag
     1801 agctgcacgc cattctgcgg cggcaggaag atttttaccc attcctgaag gacaaccggg
     1861 aaaagatcga gaagatcctg accttccgca tcccctacta cgtgggccct ctggccaggg
     1921 gaaacagcag attcgcctgg atgaccagaa agagcgagga aaccatcacc ccctggaact
     1981 tcgaggaagt ggtggacaag ggcgcttccg cccagagctt catcgagcgg atgaccaact
     2041 tcgataagaa cctgcccaac gagaaggtgc tgcccaagca cagcctgctg tacgagtact
     2101 tcaccgtgta taacgagctg accaaagtga aatacgtgac cgagggaatg agaaagcccg
     2161 ccttcctgag cggcgagcag aaaaaggcca tcgtggacct gctgttcaag accaaccgga
     2221 aagtgaccgt gaagcagctg aaagaggact acttcaagaa aatcgagtgc ttcgactccg
     2281 tggaaatctc cggcgtggaa gatcggttca acgcctccct gggcacatac cacgatctgc
     2341 tgaaaattat caaggacaag gacttcctgg acaatgagga aaacgaggac attctggaag
     2401 atatcgtgct gaccctgaca ctgtttgagg acagagagat gatcgaggaa cggctgaaaa
     2461 cctatgccca cctgttcgac gacaaagtga tgaagcagct gaagcggcgg agatacaccg
     2521 gctggggcag gctgagccgg aagctgatca acggcatccg ggacaagcag tccggcaaga
     2581 caatcctgga tttcctgaag tccgacggct tcgccaacag aaacttcatg cagctgatcc
     2641 acgacgacag cctgaccttt aaagaggaca tccagaaagc ccaggtgtcc ggccagggcg
     2701 atagcctgca cgagcacatt gccaatctgg ccggcagccc cgccattaag aagggcatcc
     2761 tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat gggccggcac aagcccgaga
     2821 acatcgtgat cgaaatggcc agagagaacc agaccaccca gaagggacag aagaacagcc
     2881 gcgagagaat gaagcggatc gaagagggca tcaaagagct gggcagccag atcctgaaag
     2941 aacaccccgt ggaaaacacc cagctgcaga acgagaagct gtacctgtac tacctgcaga
     3001 atgggcggga tatgtacgtg gaccaggaac tggacatcaa ccggctgtcc gactacgatg
     3061 tggaccatat cgtgcctcag agctttctga aggacgactc catcgacaac aaggtgctga
     3121 ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc ctccgaagag gtcgtgaaga
     3181 agatgaagaa ctactggcgg cagctgctga acgccaagct gattacccag agaaagttcg
     3241 acaatctgac caaggccgag agaggcggcc tgagcgaact ggataaggcc ggcttcatca
     3301 agagacagct ggtggaaacc cggcagatca caaagcacgt ggcacagatc ctggactccc
     3361 ggatgaacac taagtacgac gagaatgaca agctgatccg ggaagtgaaa gtgatcaccc
     3421 tgaagtccaa gctggtgtcc gatttccgga aggatttcca gttttacaaa gtgcgcgaga
     3481 tcaacaacta ccaccacgcc cacgacgcct acctgaacgc cgtcgtggga accgccctga
     3541 tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg cgactacaag gtgtacgacg
     3601 tgcggaagat gatcgccaag agcgagcagg aaatcggcaa ggctaccgcc aagtacttct
     3661 tctacagcaa catcatgaac tttttcaaga ccgagattac cctggccaac ggcgagatcc
     3721 ggaagcggcc tctgatcgag acaaacggcg aaaccgggga gatcgtgtgg gataagggcc
     3781 gggattttgc caccgtgcgg aaagtgctga gcatgcccca agtgaatatc gtgaaaaaga
     3841 ccgaggtgca gacaggcggc ttcagcaaag agtctatcct gcccaagagg aacagcgata
     3901 agctgatcgc cagaaagaag gactgggacc ctaagaagta cggcggcttc gacagcccca
     3961 ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa gggcaagtcc aagaaactga
     4021 agagtgtgaa agagctgctg gggatcacca tcatggaaag aagcagcttc gagaagaatc
     4081 ccatcgactt tctggaagcc aagggctaca aagaagtgaa aaaggacctg atcatcaagc
     4141 tgcctaagta ctccctgttc gagctggaaa acggccggaa gagaatgctg gcctctgccg
     4201 gcgaactgca gaagggaaac gaactggccc tgccctccaa atatgtgaac ttcctgtacc
     4261 tggccagcca ctatgagaag ctgaagggct cccccgagga taatgagcag aaacagctgt
     4321 ttgtggaaca gcacaagcac tacctggacg agatcatcga gcagatcagc gagttctcca
     4381 agagagtgat cctggccgac gctaatctgg acaaagtgct gtccgcctac aacaagcacc
     4441 gggataagcc catcagagag caggccgaga atatcatcca cctgtttacc ctgaccaatc
     4501 tgggagcccc tgccgccttc aagtactttg acaccaccat cgaccggaag aggtacacca
     4561 gcaccaaaga ggtgctggac gccaccctga tccaccagag catcaccggc ctgtacgaga
     4621 cacggatcga cctgtctcag ctgggaggcg acaaaaggcc ggcggccacg aaaaaggccg
     4681 gccaggcaaa aaagaaaaag taagaattcc tagagctcgc tgatcagcct cgactgtgcc
     4741 ttctagttgc cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg
     4801 tgccactccc actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag
     4861 gtgtcattct attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga
     4921 gaatagcagg catgctgggg agcggccgca ggaaccccta gtgatggagt tggccactcc
     4981 ctctctgcgc gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg
     5041 ctttgcccgg gcggcctcag tgagcgagcg agcgcgcagc tgcctgcagg ggcgcctgat
     5101 gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatacgtc aaagcaacca
     5161 tagtacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg
     5221 accgctacac ttgccagcgc cttagcgccc gctcctttcg ctttcttccc ttcctttctc
     5281 gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt agggttccga
     5341 tttagtgctt tacggcacct cgaccccaaa aaacttgatt tgggtgatgg ttcacgtagt
     5401 gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac gttctttaat
     5461 agtggactct tgttccaaac tggaacaaca ctcaactcta tctcgggcta ttcttttgat
     5521 ttataaggga ttttgccgat ttcggtctat tggttaaaaa atgagctgat ttaacaaaaa
     5581 tttaacgcga attttaacaa aatattaacg tttacaattt tatggtgcac tctcagtaca
     5641 atctgctctg atgccgcata gttaagccag ccccgacacc cgccaacacc cgctgacgcg
     5701 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     5761 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgagacg aaagggcctc
     5821 gtgatacgcc tatttttata ggttaatgtc atgataataa tggtttctta gacgtcaggt
     5881 ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta aatacattca
     5941 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     6001 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     6061 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     6121 ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt
     6181 cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta
     6241 ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat
     6301 gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga
     6361 gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca
     6421 acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact
     6481 cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc
     6541 acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact
     6601 ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt
     6661 ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt
     6721 ggaagccgcg gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt
     6781 atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata
     6841 ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata tatactttag
     6901 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     6961 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     7021 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     7081 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     7141 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     7201 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     7261 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     7321 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     7381 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     7441 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     7501 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     7561 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     7621 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     7681 cacatgtgag ggcctatttc ccatgattcc ttcatatttg catatacgat acaaggctgt
     7741 tagagagata attggaatta atttgactgt aaacacaaag atattagtac aaaatacgtg
     7801 acgtagaaag taataatttc ttgggtagtt tgcagtttta aaattatgtt ttaaaatgga
     7861 ctatcatatg cttaccgtaa cttgaaagta tttcgatttc ttggctttat atatcttgtg
     7921 gaaaggacga aacaccgagc gcactgccgc tcatcgagtt ttagagctag aaatagcaag
     7981 ttaaaataag gctagtccgt tatcaacttg aaaaagtggc accgagtcgg tgcttttttg
     8041 ttttagagct agaaatagca agttaaaata aggctagtcc gtttttagcg cgtgcgccaa
     8101 ttctgcagac aaatggctct agaggtaccc gttacataac ttacggtaaa tggcccgcct
     8161 ggctgaccgc ccaacgaccc ccgcccattg acgtcaatag taacgccaat agggactttc
     8221 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     8281 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     8341 tgtgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     8401 atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc catctccccc
     8461 ccctccccac ccccaatttt gtattta
//
