LOCUS       sequence-330055-        9650 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Plasmid encoding for 2 gRNAs targeting the human BAK gene and a CMV 
            driven nuclease Cas9 followed by self-cleaving mCherry.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9650)
  AUTHORS   Tigano M, Vargas DC, Tremblay-Belzile S, Fu Y, Sfeir A
  TITLE     Nuclear sensing of breaks in mitochondrial DNA enhances immune 
            surveillance.
  JOURNAL   Nature. 2021 Mar;591(7850):477-481. doi: 10.1038/s41586-021-03269-w.
            Epub 2021 Feb 24.
  PUBMED    33627873
REFERENCE   2  (bases 1 to 9650)
  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/s41586-021-03269-w"; journalName: "Nature"; date: 
            "2021-03"; volume: "591"; issue: "7850"; pages: "477-481"
COMMENT     Sequence Label: pSpCas9nucl-T2A-mCherry - BAKgRNA1- BAKgRNA2
FEATURES             Location/Qualifiers
     source          1..9650
                     /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)"
     misc_RNA        270..345
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        436..684
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     misc_RNA        705..780
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        871..1156
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     intron          1382..1609
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      1621..1630
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1630..1695
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1702..1722
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1747..5847
                     /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             1747..5847
                     /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             5848..5895
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             5902..5955
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             5956..6663
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     primer_bind     complement(6101..6119)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     6369..6388
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     primer_bind     complement(6691..6708)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6697..6904
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   6913..7053
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   6913..7042
                     /label=AAV2 ITR
     rep_origin      7128..7583
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7215..7234)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7425..7446
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(7600..7619)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7719..7741
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7779..7797)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7865..7969
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7970..8038,8039..8830)
                     /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(8188..8207)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      9001..9589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9490..9509
                     /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 catgaagtcg accacgaagg ttttagagct agaaatagca agttaaaata
      301 aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc ggtgcttttt tgttttagag
      361 ctagaaatag caagttaaaa taaggctagt ccgtttttag cgcgtgcgcc aattctgcag
      421 acaaatggct ctagagaggg cctatttccc atgattcctt catatttgca tatacgatac
      481 aaggctgtta gagagataat tggaattaat ttgactgtaa acacaaagat attagtacaa
      541 aatacgtgac gtagaaagta ataatttctt gggtagtttg cagttttaaa attatgtttt
      601 aaaatggact atcatatgct taccgtaact tgaaagtatt tcgatttctt ggctttatat
      661 atcttgtgga aaggacgaaa caccggccat gctggtagac gtgtgtttta gagctagaaa
      721 tagcaagtta aaataaggct agtccgttat caacttgaaa aagtggcacc gagtcggtgc
      781 ttttttgttt tagagctaga aatagcaagt taaaataagg ctagtccgtt tttagcgcgt
      841 gcgccaattc tgcagacaaa tggcggtacc cgttacataa cttacggtaa atggcccgcc
      901 tggctgaccg cccaacgacc cccgcccatt gacgtcaata gtaacgccaa tagggacttt
      961 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
     1021 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
     1081 ttgtgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
     1141 catcgctatt accatggtcg aggtgagccc cacgttctgc ttcactctcc ccatctcccc
     1201 cccctcccca cccccaattt tgtatttatt tattttttaa ttattttgtg cagcgatggg
     1261 ggcggggggg gggggggggg gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
     1321 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
     1381 gggagtcgct gcgcgctgcc ttcgccccgt gccccgctcc gccgccgcct cgcgccgccc
     1441 gccccggctc tgactgaccg cgttactccc acaggtgagc gggcgggacg gcccttctcc
     1501 tccgggctgt aattagctga gcaagaggta agggtttaag ggatggttgg ttggtggggt
     1561 attaatgttt aattacctgg agcacctgcc tgaaatcact ttttttcagg ttggaccggt
     1621 gccaccatgg actataagga ccacgacgga gactacaagg atcatgatat tgattacaaa
     1681 gacgatgacg ataagatggc cccaaagaag aagcggaagg tcggtatcca cggagtccca
     1741 gcagccgaca agaagtacag catcggcctg gacatcggca ccaactctgt gggctgggcc
     1801 gtgatcaccg acgagtacaa ggtgcccagc aagaaattca aggtgctggg caacaccgac
     1861 cggcacagca tcaagaagaa cctgatcgga gccctgctgt tcgacagcgg cgaaacagcc
     1921 gaggccaccc ggctgaagag aaccgccaga agaagataca ccagacggaa gaaccggatc
     1981 tgctatctgc aagagatctt cagcaacgag atggccaagg tggacgacag cttcttccac
     2041 agactggaag agtccttcct ggtggaagag gataagaagc acgagcggca ccccatcttc
     2101 ggcaacatcg tggacgaggt ggcctaccac gagaagtacc ccaccatcta ccacctgaga
     2161 aagaaactgg tggacagcac cgacaaggcc gacctgcggc tgatctatct ggccctggcc
     2221 cacatgatca agttccgggg ccacttcctg atcgagggcg acctgaaccc cgacaacagc
     2281 gacgtggaca agctgttcat ccagctggtg cagacctaca accagctgtt cgaggaaaac
     2341 cccatcaacg ccagcggcgt ggacgccaag gccatcctgt ctgccagact gagcaagagc
     2401 agacggctgg aaaatctgat cgcccagctg cccggcgaga agaagaatgg cctgttcgga
     2461 aacctgattg ccctgagcct gggcctgacc cccaacttca agagcaactt cgacctggcc
     2521 gaggatgcca aactgcagct gagcaaggac acctacgacg acgacctgga caacctgctg
     2581 gcccagatcg gcgaccagta cgccgacctg tttctggccg ccaagaacct gtccgacgcc
     2641 atcctgctga gcgacatcct gagagtgaac accgagatca ccaaggcccc cctgagcgcc
     2701 tctatgatca agagatacga cgagcaccac caggacctga ccctgctgaa agctctcgtg
     2761 cggcagcagc tgcctgagaa gtacaaagag attttcttcg accagagcaa gaacggctac
     2821 gccggctaca ttgacggcgg agccagccag gaagagttct acaagttcat caagcccatc
     2881 ctggaaaaga tggacggcac cgaggaactg ctcgtgaagc tgaacagaga ggacctgctg
     2941 cggaagcagc ggaccttcga caacggcagc atcccccacc agatccacct gggagagctg
     3001 cacgccattc tgcggcggca ggaagatttt tacccattcc tgaaggacaa ccgggaaaag
     3061 atcgagaaga tcctgacctt ccgcatcccc tactacgtgg gccctctggc caggggaaac
     3121 agcagattcg cctggatgac cagaaagagc gaggaaacca tcaccccctg gaacttcgag
     3181 gaagtggtgg acaagggcgc ttccgcccag agcttcatcg agcggatgac caacttcgat
     3241 aagaacctgc ccaacgagaa ggtgctgccc aagcacagcc tgctgtacga gtacttcacc
     3301 gtgtataacg agctgaccaa agtgaaatac gtgaccgagg gaatgagaaa gcccgccttc
     3361 ctgagcggcg agcagaaaaa ggccatcgtg gacctgctgt tcaagaccaa ccggaaagtg
     3421 accgtgaagc agctgaaaga ggactacttc aagaaaatcg agtgcttcga ctccgtggaa
     3481 atctccggcg tggaagatcg gttcaacgcc tccctgggca cataccacga tctgctgaaa
     3541 attatcaagg acaaggactt cctggacaat gaggaaaacg aggacattct ggaagatatc
     3601 gtgctgaccc tgacactgtt tgaggacaga gagatgatcg aggaacggct gaaaacctat
     3661 gcccacctgt tcgacgacaa agtgatgaag cagctgaagc ggcggagata caccggctgg
     3721 ggcaggctga gccggaagct gatcaacggc atccgggaca agcagtccgg caagacaatc
     3781 ctggatttcc tgaagtccga cggcttcgcc aacagaaact tcatgcagct gatccacgac
     3841 gacagcctga cctttaaaga ggacatccag aaagcccagg tgtccggcca gggcgatagc
     3901 ctgcacgagc acattgccaa tctggccggc agccccgcca ttaagaaggg catcctgcag
     3961 acagtgaagg tggtggacga gctcgtgaaa gtgatgggcc ggcacaagcc cgagaacatc
     4021 gtgatcgaaa tggccagaga gaaccagacc acccagaagg gacagaagaa cagccgcgag
     4081 agaatgaagc ggatcgaaga gggcatcaaa gagctgggca gccagatcct gaaagaacac
     4141 cccgtggaaa acacccagct gcagaacgag aagctgtacc tgtactacct gcagaatggg
     4201 cgggatatgt acgtggacca ggaactggac atcaaccggc tgtccgacta cgatgtggac
     4261 catatcgtgc ctcagagctt tctgaaggac gactccatcg acaacaaggt gctgaccaga
     4321 agcgacaaga accggggcaa gagcgacaac gtgccctccg aagaggtcgt gaagaagatg
     4381 aagaactact ggcggcagct gctgaacgcc aagctgatta cccagagaaa gttcgacaat
     4441 ctgaccaagg ccgagagagg cggcctgagc gaactggata aggccggctt catcaagaga
     4501 cagctggtgg aaacccggca gatcacaaag cacgtggcac agatcctgga ctcccggatg
     4561 aacactaagt acgacgagaa tgacaagctg atccgggaag tgaaagtgat caccctgaag
     4621 tccaagctgg tgtccgattt ccggaaggat ttccagtttt acaaagtgcg cgagatcaac
     4681 aactaccacc acgcccacga cgcctacctg aacgccgtcg tgggaaccgc cctgatcaaa
     4741 aagtacccta agctggaaag cgagttcgtg tacggcgact acaaggtgta cgacgtgcgg
     4801 aagatgatcg ccaagagcga gcaggaaatc ggcaaggcta ccgccaagta cttcttctac
     4861 agcaacatca tgaacttttt caagaccgag attaccctgg ccaacggcga gatccggaag
     4921 cggcctctga tcgagacaaa cggcgaaacc ggggagatcg tgtgggataa gggccgggat
     4981 tttgccaccg tgcggaaagt gctgagcatg ccccaagtga atatcgtgaa aaagaccgag
     5041 gtgcagacag gcggcttcag caaagagtct atcctgccca agaggaacag cgataagctg
     5101 atcgccagaa agaaggactg ggaccctaag aagtacggcg gcttcgacag ccccaccgtg
     5161 gcctattctg tgctggtggt ggccaaagtg gaaaagggca agtccaagaa actgaagagt
     5221 gtgaaagagc tgctggggat caccatcatg gaaagaagca gcttcgagaa gaatcccatc
     5281 gactttctgg aagccaaggg ctacaaagaa gtgaaaaagg acctgatcat caagctgcct
     5341 aagtactccc tgttcgagct ggaaaacggc cggaagagaa tgctggcctc tgccggcgaa
     5401 ctgcagaagg gaaacgaact ggccctgccc tccaaatatg tgaacttcct gtacctggcc
     5461 agccactatg agaagctgaa gggctccccc gaggataatg agcagaaaca gctgtttgtg
     5521 gaacagcaca agcactacct ggacgagatc atcgagcaga tcagcgagtt ctccaagaga
     5581 gtgatcctgg ccgacgctaa tctggacaaa gtgctgtccg cctacaacaa gcaccgggat
     5641 aagcccatca gagagcaggc cgagaatatc atccacctgt ttaccctgac caatctggga
     5701 gcccctgccg ccttcaagta ctttgacacc accatcgacc ggaagaggta caccagcacc
     5761 aaagaggtgc tggacgccac cctgatccac cagagcatca ccggcctgta cgagacacgg
     5821 atcgacctgt ctcagctggg aggcgacaaa aggccggcgg ccacgaaaaa ggccggccag
     5881 gcaaaaaaga aaaaggaatt cgaaggaagg ggctctttgc ttacttgtgg agatgttgag
     5941 gaaaatccag gacccatggt gagcaagggc gaggaggata acatggccat catcaaggag
     6001 ttcatgcgct tcaaggtgca catggagggc tccgtgaacg gccacgagtt cgagatcgag
     6061 ggcgagggcg agggccgccc ctacgagggc acccagaccg ccaagctgaa ggtgaccaag
     6121 ggtggccccc tgcccttcgc ctgggacatc ctgtcccctc agttcatgta cggctccaag
     6181 gcctacgtga agcaccccgc cgacatcccc gactacttga agctgtcctt ccccgagggc
     6241 ttcaagtggg agcgcgtgat gaacttcgag gacggcggcg tggtgaccgt gacccaggac
     6301 tcctccctgc aggacggcga gttcatctac aaggtgaagc tgcgcggcac caacttcccc
     6361 tccgacggcc ccgtaatgca gaagaagacc atgggctggg aggcctcctc cgagcggatg
     6421 taccccgagg acggcgccct gaagggcgag atcaagcaga ggctgaagct gaaggacggc
     6481 ggccactacg acgctgaggt caagaccacc tacaaggcca agaagcccgt gcagctgccc
     6541 ggcgcctaca acgtcaacat caagttggac atcacctccc acaacgagga ctacaccatc
     6601 gtggaacagt acgaacgcgc cgagggccgc cactccaccg gcggcatgga cgagctgtac
     6661 aaggaattct aactagagct cgctgatcag cctcgactgt gccttctagt tgccagccat
     6721 ctgttgtttg cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc
     6781 tttcctaata aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg
     6841 ggggtggggt ggggcaggac agcaaggggg aggattggga agagaatagc aggcatgctg
     6901 gggagcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct
     6961 cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct
     7021 cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta
     7081 cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag
     7141 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     7201 cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     7261 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     7321 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     7381 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     7441 aactggaaca acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc
     7501 gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     7561 caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc
     7621 atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct
     7681 gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag
     7741 gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt
     7801 ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa
     7861 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     7921 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     7981 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     8041 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     8101 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     8161 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     8221 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     8281 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     8341 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     8401 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     8461 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     8521 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     8581 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     8641 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtggaagcc gcggtatcat
     8701 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     8761 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     8821 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     8881 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     8941 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     9001 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     9061 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     9121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     9181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     9241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     9301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     9361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     9421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     9481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     9541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     9601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
