LOCUS       sequence-455792-        8974 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SpCas9, the PITCh gRNA, and a sgRNA targeting the 
            C-terminus of TOMM20 for knock-in..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8974)
  AUTHORS   Philip R, Sharma A, Matellan L, Erpf AC, Hsu WH, Tkach JM, Wyatt 
            HDM, Pelletier L
  TITLE     qTAG: an adaptable plasmid scaffold for CRISPR-based endogenous 
            tagging.
  JOURNAL   EMBO J. 2024 Dec 12. doi: 10.1038/s44318-024-00337-5.
  PUBMED    39668248
REFERENCE   2  (bases 1 to 8974)
  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: "EMBO J. 
            2024 Dec 12. doi: 10.1038/s44318-024-00337-5."
COMMENT     Sequence Label: pX330-PITCh-TOMM20
FEATURES             Location/Qualifiers
     source          1..8974
                     /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          590..817
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      829..838
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             838..903
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             910..930
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             955..5055
                     /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             955..5055
                     /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             5056..5103
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5131..5148)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5137..5344
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5353..5493
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5353..5482
                     /label=AAV2 ITR
     rep_origin      5568..6023
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5655..5674)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5865..5886
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6040..6059)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6159..6181
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6219..6237)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6305..6409
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6410..6478,6479..7270)
                     /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(6628..6647)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7441..8029
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7930..7949
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        8091..8339
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        8361..8436
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        8534..8782
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        8803..8878
                     /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 cgcgccgggg
      421 gggggggggg gggggggggg gggggggggg gggggggggg gggggggggg ggggggcgag
      481 gcggagaggt gcggcggcag ccaatcagag cggcgcgctc cgaaagtttc cttttatggc
      541 gaggcggcgg cggcggcggc cctataaaaa gcgaagcgcg cggcgggcgg gagtcgctgc
      601 gcgctgcctt cgccccgtgc cccgctccgc cgccgcctcg cgccgcccgc cccggctctg
      661 actgaccgcg ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa
      721 ttagctgagc aagaggtaag ggtttaaggg atggttggtt ggtggggtat taatgtttaa
      781 ttacctggag cacctgcctg aaatcacttt ttttcaggtt ggaccggtgc caccatggac
      841 tataaggacc acgacggaga ctacaaggat catgatattg attacaaaga cgatgacgat
      901 aagatggccc caaagaagaa gcggaaggtc ggtatccacg gagtcccagc agccgacaag
      961 aagtacagca tcggcctgga catcggcacc aactctgtgg gctgggccgt gatcaccgac
     1021 gagtacaagg tgcccagcaa gaaattcaag gtgctgggca acaccgaccg gcacagcatc
     1081 aagaagaacc tgatcggagc cctgctgttc gacagcggcg aaacagccga ggccacccgg
     1141 ctgaagagaa ccgccagaag aagatacacc agacggaaga accggatctg ctatctgcaa
     1201 gagatcttca gcaacgagat ggccaaggtg gacgacagct tcttccacag actggaagag
     1261 tccttcctgg tggaagagga taagaagcac gagcggcacc ccatcttcgg caacatcgtg
     1321 gacgaggtgg cctaccacga gaagtacccc accatctacc acctgagaaa gaaactggtg
     1381 gacagcaccg acaaggccga cctgcggctg atctatctgg ccctggccca catgatcaag
     1441 ttccggggcc acttcctgat cgagggcgac ctgaaccccg acaacagcga cgtggacaag
     1501 ctgttcatcc agctggtgca gacctacaac cagctgttcg aggaaaaccc catcaacgcc
     1561 agcggcgtgg acgccaaggc catcctgtct gccagactga gcaagagcag acggctggaa
     1621 aatctgatcg cccagctgcc cggcgagaag aagaatggcc tgttcggaaa cctgattgcc
     1681 ctgagcctgg gcctgacccc caacttcaag agcaacttcg acctggccga ggatgccaaa
     1741 ctgcagctga gcaaggacac ctacgacgac gacctggaca acctgctggc ccagatcggc
     1801 gaccagtacg ccgacctgtt tctggccgcc aagaacctgt ccgacgccat cctgctgagc
     1861 gacatcctga gagtgaacac cgagatcacc aaggcccccc tgagcgcctc tatgatcaag
     1921 agatacgacg agcaccacca ggacctgacc ctgctgaaag ctctcgtgcg gcagcagctg
     1981 cctgagaagt acaaagagat tttcttcgac cagagcaaga acggctacgc cggctacatt
     2041 gacggcggag ccagccagga agagttctac aagttcatca agcccatcct ggaaaagatg
     2101 gacggcaccg aggaactgct cgtgaagctg aacagagagg acctgctgcg gaagcagcgg
     2161 accttcgaca acggcagcat cccccaccag atccacctgg gagagctgca cgccattctg
     2221 cggcggcagg aagattttta cccattcctg aaggacaacc gggaaaagat cgagaagatc
     2281 ctgaccttcc gcatccccta ctacgtgggc cctctggcca ggggaaacag cagattcgcc
     2341 tggatgacca gaaagagcga ggaaaccatc accccctgga acttcgagga agtggtggac
     2401 aagggcgctt ccgcccagag cttcatcgag cggatgacca acttcgataa gaacctgccc
     2461 aacgagaagg tgctgcccaa gcacagcctg ctgtacgagt acttcaccgt gtataacgag
     2521 ctgaccaaag tgaaatacgt gaccgaggga atgagaaagc ccgccttcct gagcggcgag
     2581 cagaaaaagg ccatcgtgga cctgctgttc aagaccaacc ggaaagtgac cgtgaagcag
     2641 ctgaaagagg actacttcaa gaaaatcgag tgcttcgact ccgtggaaat ctccggcgtg
     2701 gaagatcggt tcaacgcctc cctgggcaca taccacgatc tgctgaaaat tatcaaggac
     2761 aaggacttcc tggacaatga ggaaaacgag gacattctgg aagatatcgt gctgaccctg
     2821 acactgtttg aggacagaga gatgatcgag gaacggctga aaacctatgc ccacctgttc
     2881 gacgacaaag tgatgaagca gctgaagcgg cggagataca ccggctgggg caggctgagc
     2941 cggaagctga tcaacggcat ccgggacaag cagtccggca agacaatcct ggatttcctg
     3001 aagtccgacg gcttcgccaa cagaaacttc atgcagctga tccacgacga cagcctgacc
     3061 tttaaagagg acatccagaa agcccaggtg tccggccagg gcgatagcct gcacgagcac
     3121 attgccaatc tggccggcag ccccgccatt aagaagggca tcctgcagac agtgaaggtg
     3181 gtggacgagc tcgtgaaagt gatgggccgg cacaagcccg agaacatcgt gatcgaaatg
     3241 gccagagaga accagaccac ccagaaggga cagaagaaca gccgcgagag aatgaagcgg
     3301 atcgaagagg gcatcaaaga gctgggcagc cagatcctga aagaacaccc cgtggaaaac
     3361 acccagctgc agaacgagaa gctgtacctg tactacctgc agaatgggcg ggatatgtac
     3421 gtggaccagg aactggacat caaccggctg tccgactacg atgtggacca tatcgtgcct
     3481 cagagctttc tgaaggacga ctccatcgac aacaaggtgc tgaccagaag cgacaagaac
     3541 cggggcaaga gcgacaacgt gccctccgaa gaggtcgtga agaagatgaa gaactactgg
     3601 cggcagctgc tgaacgccaa gctgattacc cagagaaagt tcgacaatct gaccaaggcc
     3661 gagagaggcg gcctgagcga actggataag gccggcttca tcaagagaca gctggtggaa
     3721 acccggcaga tcacaaagca cgtggcacag atcctggact cccggatgaa cactaagtac
     3781 gacgagaatg acaagctgat ccgggaagtg aaagtgatca ccctgaagtc caagctggtg
     3841 tccgatttcc ggaaggattt ccagttttac aaagtgcgcg agatcaacaa ctaccaccac
     3901 gcccacgacg cctacctgaa cgccgtcgtg ggaaccgccc tgatcaaaaa gtaccctaag
     3961 ctggaaagcg agttcgtgta cggcgactac aaggtgtacg acgtgcggaa gatgatcgcc
     4021 aagagcgagc aggaaatcgg caaggctacc gccaagtact tcttctacag caacatcatg
     4081 aactttttca agaccgagat taccctggcc aacggcgaga tccggaagcg gcctctgatc
     4141 gagacaaacg gcgaaaccgg ggagatcgtg tgggataagg gccgggattt tgccaccgtg
     4201 cggaaagtgc tgagcatgcc ccaagtgaat atcgtgaaaa agaccgaggt gcagacaggc
     4261 ggcttcagca aagagtctat cctgcccaag aggaacagcg ataagctgat cgccagaaag
     4321 aaggactggg accctaagaa gtacggcggc ttcgacagcc ccaccgtggc ctattctgtg
     4381 ctggtggtgg ccaaagtgga aaagggcaag tccaagaaac tgaagagtgt gaaagagctg
     4441 ctggggatca ccatcatgga aagaagcagc ttcgagaaga atcccatcga ctttctggaa
     4501 gccaagggct acaaagaagt gaaaaaggac ctgatcatca agctgcctaa gtactccctg
     4561 ttcgagctgg aaaacggccg gaagagaatg ctggcctctg ccggcgaact gcagaaggga
     4621 aacgaactgg ccctgccctc caaatatgtg aacttcctgt acctggccag ccactatgag
     4681 aagctgaagg gctcccccga ggataatgag cagaaacagc tgtttgtgga acagcacaag
     4741 cactacctgg acgagatcat cgagcagatc agcgagttct ccaagagagt gatcctggcc
     4801 gacgctaatc tggacaaagt gctgtccgcc tacaacaagc accgggataa gcccatcaga
     4861 gagcaggccg agaatatcat ccacctgttt accctgacca atctgggagc ccctgccgcc
     4921 ttcaagtact ttgacaccac catcgaccgg aagaggtaca ccagcaccaa agaggtgctg
     4981 gacgccaccc tgatccacca gagcatcacc ggcctgtacg agacacggat cgacctgtct
     5041 cagctgggag gcgacaaaag gccggcggcc acgaaaaagg ccggccaggc aaaaaagaaa
     5101 aagtaagaat tcctagagct cgctgatcag cctcgactgt gccttctagt tgccagccat
     5161 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     5221 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     5281 ggggtggggt ggggcaggac agcaaggggg aggattggga agagaatagc aggcatgctg
     5341 gggagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct
     5401 cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct
     5461 cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta
     5521 cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag
     5581 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     5641 cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     5701 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     5761 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     5821 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     5881 aactggaaca acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc
     5941 gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     6001 caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc
     6061 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     6121 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     6181 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     6241 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     6301 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     6361 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     6421 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     6481 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     6541 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     6601 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     6661 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     6721 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     6781 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     6841 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     6901 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     6961 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     7021 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     7081 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtggaagcc gcggtatcat
     7141 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     7201 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     7261 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     7321 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     7381 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     7441 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     7501 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     7561 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     7621 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     7681 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     7741 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     7801 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     7861 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     7921 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     7981 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     8041 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt gagggcctat
     8101 ttcccatgat tccttcatat ttgcatatac gatacaaggc tgttagagag ataattggaa
     8161 ttaatttgac tgtaaacaca aagatattag tacaaaatac gtgacgtaga aagtaataat
     8221 ttcttgggta gtttgcagtt ttaaaattat gttttaaaat ggactatcat atgcttaccg
     8281 taacttgaaa gtatttcgat ttcttggctt tatatatctt gtggaaagga cgaaacaccg
     8341 gagcttggct gaagatgatg gttttagagc tagaaatagc aagttaaaat aaggctagtc
     8401 cgttatcaac ttgaaaaagt ggcaccgagt cggtgctttt ttgttttaga gctagaaata
     8461 gcaagttaaa ataaggctag tccgttttta gcgcgtgcgc caattctgca gacaaatggc
     8521 tctagaggca tgtgagggcc tatttcccat gattccttca tatttgcata tacgatacaa
     8581 ggctgttaga gagataattg gaattaattt gactgtaaac acaaagatat tagtacaaaa
     8641 tacgtgacgt agaaagtaat aatttcttgg gtagtttgca gttttaaaat tatgttttaa
     8701 aatggactat catatgctta ccgtaacttg aaagtatttc gatttcttgg ctttatatat
     8761 cttgtggaaa ggacgaaaca ccgcatcgta cgcgtacgtg ttgttttaga gctagaaata
     8821 gcaagttaaa ataaggctag tccgttatca acttgaaaaa gtggcaccga gtcggtgctt
     8881 ttttgtttta gagctagaaa tagcaagtta aaataaggct agtccgtttt tagcgcgtgc
     8941 gccaattctg cagacaaatg gctctagagg tacc
//
