LOCUS       sequence-416183-        8950 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SpCas9, the PITCh gRNA, and a sgRNA targeting the 
            C-terminus of TUBB4B for knock-in..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8950)
  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 8950)
  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-TUBB4B
FEATURES             Location/Qualifiers
     source          1..8950
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        8..293
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        295..572
                     /label=chicken beta-actin promoter
     intron          573..800
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      812..821
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             821..886
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             893..913
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             938..5038
                     /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             938..5038
                     /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             5039..5086
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     primer_bind     complement(5114..5131)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    5120..5327
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5336..5476
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5336..5465
                     /label=AAV2 ITR
     rep_origin      5551..6006
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5638..5657)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5848..5869
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6023..6042)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6142..6164
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6202..6220)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6288..6392
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6393..6461,6462..7253)
                     /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(6611..6630)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7424..8012
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7913..7932
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        8074..8322
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        8344..8419
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        8517..8765
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        8786..8861
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
ORIGIN
        1 aggtacccgt tacataactt acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc
       61 gcccattgac gtcaatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt
      121 atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc
      181 ctattgacgt caatgacggt aaatggcccg cctggcattg tgcccagtac atgaccttat
      241 gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc atggtcgagg
      301 tgagccccac gttctgcttc actctcccca tctccccccc ctccccaccc ccaattttgt
      361 atttatttat tttttaatta ttttgtgcag cgatgggggc gggggggggg ggggggcgcg
      421 cgccgggggg gggggggggg gggggggggg gggggggggc gaggcggaga ggtgcggcgg
      481 cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg cggcggcggc
      541 ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgcgctgc cttcgccccg
      601 tgccccgctc cgccgccgcc tcgcgccgcc cgccccggct ctgactgacc gcgttactcc
      661 cacaggtgag cgggcgggac ggcccttctc ctccgggctg taattagctg agcaagaggt
      721 aagggtttaa gggatggttg gttggtgggg tattaatgtt taattacctg gagcacctgc
      781 ctgaaatcac tttttttcag gttggaccgg tgccaccatg gactataagg accacgacgg
      841 agactacaag gatcatgata ttgattacaa agacgatgac gataagatgg ccccaaagaa
      901 gaagcggaag gtcggtatcc acggagtccc agcagccgac aagaagtaca gcatcggcct
      961 ggacatcggc accaactctg tgggctgggc cgtgatcacc gacgagtaca aggtgcccag
     1021 caagaaattc aaggtgctgg gcaacaccga ccggcacagc atcaagaaga acctgatcgg
     1081 agccctgctg ttcgacagcg gcgaaacagc cgaggccacc cggctgaaga gaaccgccag
     1141 aagaagatac accagacgga agaaccggat ctgctatctg caagagatct tcagcaacga
     1201 gatggccaag gtggacgaca gcttcttcca cagactggaa gagtccttcc tggtggaaga
     1261 ggataagaag cacgagcggc accccatctt cggcaacatc gtggacgagg tggcctacca
     1321 cgagaagtac cccaccatct accacctgag aaagaaactg gtggacagca ccgacaaggc
     1381 cgacctgcgg ctgatctatc tggccctggc ccacatgatc aagttccggg gccacttcct
     1441 gatcgagggc gacctgaacc ccgacaacag cgacgtggac aagctgttca tccagctggt
     1501 gcagacctac aaccagctgt tcgaggaaaa ccccatcaac gccagcggcg tggacgccaa
     1561 ggccatcctg tctgccagac tgagcaagag cagacggctg gaaaatctga tcgcccagct
     1621 gcccggcgag aagaagaatg gcctgttcgg aaacctgatt gccctgagcc tgggcctgac
     1681 ccccaacttc aagagcaact tcgacctggc cgaggatgcc aaactgcagc tgagcaagga
     1741 cacctacgac gacgacctgg acaacctgct ggcccagatc ggcgaccagt acgccgacct
     1801 gtttctggcc gccaagaacc tgtccgacgc catcctgctg agcgacatcc tgagagtgaa
     1861 caccgagatc accaaggccc ccctgagcgc ctctatgatc aagagatacg acgagcacca
     1921 ccaggacctg accctgctga aagctctcgt gcggcagcag ctgcctgaga agtacaaaga
     1981 gattttcttc gaccagagca agaacggcta cgccggctac attgacggcg gagccagcca
     2041 ggaagagttc tacaagttca tcaagcccat cctggaaaag atggacggca ccgaggaact
     2101 gctcgtgaag ctgaacagag aggacctgct gcggaagcag cggaccttcg acaacggcag
     2161 catcccccac cagatccacc tgggagagct gcacgccatt ctgcggcggc aggaagattt
     2221 ttacccattc ctgaaggaca accgggaaaa gatcgagaag atcctgacct tccgcatccc
     2281 ctactacgtg ggccctctgg ccaggggaaa cagcagattc gcctggatga ccagaaagag
     2341 cgaggaaacc atcaccccct ggaacttcga ggaagtggtg gacaagggcg cttccgccca
     2401 gagcttcatc gagcggatga ccaacttcga taagaacctg cccaacgaga aggtgctgcc
     2461 caagcacagc ctgctgtacg agtacttcac cgtgtataac gagctgacca aagtgaaata
     2521 cgtgaccgag ggaatgagaa agcccgcctt cctgagcggc gagcagaaaa aggccatcgt
     2581 ggacctgctg ttcaagacca accggaaagt gaccgtgaag cagctgaaag aggactactt
     2641 caagaaaatc gagtgcttcg actccgtgga aatctccggc gtggaagatc ggttcaacgc
     2701 ctccctgggc acataccacg atctgctgaa aattatcaag gacaaggact tcctggacaa
     2761 tgaggaaaac gaggacattc tggaagatat cgtgctgacc ctgacactgt ttgaggacag
     2821 agagatgatc gaggaacggc tgaaaaccta tgcccacctg ttcgacgaca aagtgatgaa
     2881 gcagctgaag cggcggagat acaccggctg gggcaggctg agccggaagc tgatcaacgg
     2941 catccgggac aagcagtccg gcaagacaat cctggatttc ctgaagtccg acggcttcgc
     3001 caacagaaac ttcatgcagc tgatccacga cgacagcctg acctttaaag aggacatcca
     3061 gaaagcccag gtgtccggcc agggcgatag cctgcacgag cacattgcca atctggccgg
     3121 cagccccgcc attaagaagg gcatcctgca gacagtgaag gtggtggacg agctcgtgaa
     3181 agtgatgggc cggcacaagc ccgagaacat cgtgatcgaa atggccagag agaaccagac
     3241 cacccagaag ggacagaaga acagccgcga gagaatgaag cggatcgaag agggcatcaa
     3301 agagctgggc agccagatcc tgaaagaaca ccccgtggaa aacacccagc tgcagaacga
     3361 gaagctgtac ctgtactacc tgcagaatgg gcgggatatg tacgtggacc aggaactgga
     3421 catcaaccgg ctgtccgact acgatgtgga ccatatcgtg cctcagagct ttctgaagga
     3481 cgactccatc gacaacaagg tgctgaccag aagcgacaag aaccggggca agagcgacaa
     3541 cgtgccctcc gaagaggtcg tgaagaagat gaagaactac tggcggcagc tgctgaacgc
     3601 caagctgatt acccagagaa agttcgacaa tctgaccaag gccgagagag gcggcctgag
     3661 cgaactggat aaggccggct tcatcaagag acagctggtg gaaacccggc agatcacaaa
     3721 gcacgtggca cagatcctgg actcccggat gaacactaag tacgacgaga atgacaagct
     3781 gatccgggaa gtgaaagtga tcaccctgaa gtccaagctg gtgtccgatt tccggaagga
     3841 tttccagttt tacaaagtgc gcgagatcaa caactaccac cacgcccacg acgcctacct
     3901 gaacgccgtc gtgggaaccg ccctgatcaa aaagtaccct aagctggaaa gcgagttcgt
     3961 gtacggcgac tacaaggtgt acgacgtgcg gaagatgatc gccaagagcg agcaggaaat
     4021 cggcaaggct accgccaagt acttcttcta cagcaacatc atgaactttt tcaagaccga
     4081 gattaccctg gccaacggcg agatccggaa gcggcctctg atcgagacaa acggcgaaac
     4141 cggggagatc gtgtgggata agggccggga ttttgccacc gtgcggaaag tgctgagcat
     4201 gccccaagtg aatatcgtga aaaagaccga ggtgcagaca ggcggcttca gcaaagagtc
     4261 tatcctgccc aagaggaaca gcgataagct gatcgccaga aagaaggact gggaccctaa
     4321 gaagtacggc ggcttcgaca gccccaccgt ggcctattct gtgctggtgg tggccaaagt
     4381 ggaaaagggc aagtccaaga aactgaagag tgtgaaagag ctgctgggga tcaccatcat
     4441 ggaaagaagc agcttcgaga agaatcccat cgactttctg gaagccaagg gctacaaaga
     4501 agtgaaaaag gacctgatca tcaagctgcc taagtactcc ctgttcgagc tggaaaacgg
     4561 ccggaagaga atgctggcct ctgccggcga actgcagaag ggaaacgaac tggccctgcc
     4621 ctccaaatat gtgaacttcc tgtacctggc cagccactat gagaagctga agggctcccc
     4681 cgaggataat gagcagaaac agctgtttgt ggaacagcac aagcactacc tggacgagat
     4741 catcgagcag atcagcgagt tctccaagag agtgatcctg gccgacgcta atctggacaa
     4801 agtgctgtcc gcctacaaca agcaccggga taagcccatc agagagcagg ccgagaatat
     4861 catccacctg tttaccctga ccaatctggg agcccctgcc gccttcaagt actttgacac
     4921 caccatcgac cggaagaggt acaccagcac caaagaggtg ctggacgcca ccctgatcca
     4981 ccagagcatc accggcctgt acgagacacg gatcgacctg tctcagctgg gaggcgacaa
     5041 aaggccggcg gccacgaaaa aggccggcca ggcaaaaaag aaaaagtaag aattcctaga
     5101 gctcgctgat cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc
     5161 cccgtgcctt ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag
     5221 gaaattgcat cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag
     5281 gacagcaagg gggaggattg ggaagagaat agcaggcatg ctggggagcg gccgcaggaa
     5341 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     5401 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     5461 cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     5521 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg
     5581 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc
     5641 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5701 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5761 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     5821 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5881 actctatctc gggctattct tttgatttat aagggatttt gccgatttcg gtctattggt
     5941 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     6001 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     6061 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     6121 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     6181 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     6241 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     6301 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     6361 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     6421 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     6481 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     6541 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     6601 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     6661 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     6721 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     6781 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     6841 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6901 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     6961 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     7021 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     7081 ctgataaatc tggagccggt gagcgtggaa gccgcggtat cattgcagca ctggggccag
     7141 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     7201 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     7261 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     7321 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     7381 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     7441 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     7501 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     7561 caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     7621 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     7681 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     7741 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     7801 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7861 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7921 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7981 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     8041 tcctggcctt ttgctggcct tttgctcaca tgtgagggcc tatttcccat gattccttca
     8101 tatttgcata tacgatacaa ggctgttaga gagataattg gaattaattt gactgtaaac
     8161 acaaagatat tagtacaaaa tacgtgacgt agaaagtaat aatttcttgg gtagtttgca
     8221 gttttaaaat tatgttttaa aatggactat catatgctta ccgtaacttg aaagtatttc
     8281 gatttcttgg ctttatatat cttgtggaaa ggacgaaaca ccgcctagag ccttcagtca
     8341 ctggttttag agctagaaat agcaagttaa aataaggcta gtccgttatc aacttgaaaa
     8401 agtggcaccg agtcggtgct tttttgtttt agagctagaa atagcaagtt aaaataaggc
     8461 tagtccgttt ttagcgcgtg cgccaattct gcagacaaat ggctctagag gcatgtgagg
     8521 gcctatttcc catgattcct tcatatttgc atatacgata caaggctgtt agagagataa
     8581 ttggaattaa tttgactgta aacacaaaga tattagtaca aaatacgtga cgtagaaagt
     8641 aataatttct tgggtagttt gcagttttaa aattatgttt taaaatggac tatcatatgc
     8701 ttaccgtaac ttgaaagtat ttcgatttct tggctttata tatcttgtgg aaaggacgaa
     8761 acaccgcatc gtacgcgtac gtgttgtttt agagctagaa atagcaagtt aaaataaggc
     8821 tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg cttttttgtt ttagagctag
     8881 aaatagcaag ttaaaataag gctagtccgt ttttagcgcg tgcgccaatt ctgcagacaa
     8941 atggctctag
//
