LOCUS       sequence-314868-        9278 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Cas9-2A-mCherry and a sgRNA excising EGFP flanked by a 
            splice acceptor and a splice donor from the 
            Intron-Tagging-EGFP-Donor plasmid..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9278)
  AUTHORS   Reicher A, Koren A, Kubicek S
  TITLE     Pooled protein tagging, cellular imaging, and in situ sequencing for
            monitoring drug action in real time.
  JOURNAL   Genome Res. 2020 Dec;30(12):1846-1855. doi: 10.1101/gr.261503.120. 
            Epub 2020 Nov 17.
  PUBMED    33203764
REFERENCE   2  (bases 1 to 9278)
  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: 
            "10.1101/gr.261503.120"; journalName: "Genome Res"; date: "2020-12";
            volume: "30"; issue: "12"; pages: "1846-1855"
COMMENT     Sequence Label: Intron-Tagging-pX330-Cas9-mCherry
FEATURES             Location/Qualifiers
     source          1..9278
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             3773..3820
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     CDS             3824..3877
                     /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             3884..4594
                     /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(4029..4047)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     complement(4619..4636)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4625..4832
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4841..4981
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4841..4970
                     /label=AAV2 ITR
     rep_origin      5056..5511
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5143..5162)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5353..5374
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5528..5547)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5647..5669
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5707..5725)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5793..5897
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5898..5966,5967..6758)
                     /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(6116..6135)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6929..7517
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7418..7437
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        7579..7827
                     /label=U6 promoter
                     /note="RNA polymerase III promoter for human U6 snRNA 
                     (Domitrovich & Kunkel, 2003)"
     primer_bind     7579..7599
                     /label=hU6-F
                     /note="Human U6 promoter, forward primer"
     primer_bind     7750..7769
                     /label=LKO.1 5'
                     /note="Human U6 promoter, forward primer"
     misc_RNA        7848..7923
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     enhancer        8020..8305
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer; 
                     contains an 18-bp deletion relative to the standard CMV 
                     enhancer"
     promoter        8307..8584
                     /label=chicken beta-actin promoter
     intron          8585..8812
                     /label=hybrid intron
                     /note="hybrid between chicken beta-actin (CBA) and minute 
                     virus of mice (MMV) introns (Gray et al., 2011)"
     regulatory      8824..8833
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             8833..8898
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             8905..8925
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(8950..9278,1..3772)
                     /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             join(8950..9278,1..3772)
                     /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"
ORIGIN
        1 gaagcacgag cggcacccca tcttcggcaa catcgtggac gaggtggcct accacgagaa
       61 gtaccccacc atctaccacc tgagaaagaa actggtggac agcaccgaca aggccgacct
      121 gcggctgatc tatctggccc tggcccacat gatcaagttc cggggccact tcctgatcga
      181 gggcgacctg aaccccgaca acagcgacgt ggacaagctg ttcatccagc tggtgcagac
      241 ctacaaccag ctgttcgagg aaaaccccat caacgccagc ggcgtggacg ccaaggccat
      301 cctgtctgcc agactgagca agagcagacg gctggaaaat ctgatcgccc agctgcccgg
      361 cgagaagaag aatggcctgt tcggaaacct gattgccctg agcctgggcc tgacccccaa
      421 cttcaagagc aacttcgacc tggccgagga tgccaaactg cagctgagca aggacaccta
      481 cgacgacgac ctggacaacc tgctggccca gatcggcgac cagtacgccg acctgtttct
      541 ggccgccaag aacctgtccg acgccatcct gctgagcgac atcctgagag tgaacaccga
      601 gatcaccaag gcccccctga gcgcctctat gatcaagaga tacgacgagc accaccagga
      661 cctgaccctg ctgaaagctc tcgtgcggca gcagctgcct gagaagtaca aagagatttt
      721 cttcgaccag agcaagaacg gctacgccgg ctacattgac ggcggagcca gccaggaaga
      781 gttctacaag ttcatcaagc ccatcctgga aaagatggac ggcaccgagg aactgctcgt
      841 gaagctgaac agagaggacc tgctgcggaa gcagcggacc ttcgacaacg gcagcatccc
      901 ccaccagatc cacctgggag agctgcacgc cattctgcgg cggcaggaag atttttaccc
      961 attcctgaag gacaaccggg aaaagatcga gaagatcctg accttccgca tcccctacta
     1021 cgtgggccct ctggccaggg gaaacagcag attcgcctgg atgaccagaa agagcgagga
     1081 aaccatcacc ccctggaact tcgaggaagt ggtggacaag ggcgcttccg cccagagctt
     1141 catcgagcgg atgaccaact tcgataagaa cctgcccaac gagaaggtgc tgcccaagca
     1201 cagcctgctg tacgagtact tcaccgtgta taacgagctg accaaagtga aatacgtgac
     1261 cgagggaatg agaaagcccg ccttcctgag cggcgagcag aaaaaggcca tcgtggacct
     1321 gctgttcaag accaaccgga aagtgaccgt gaagcagctg aaagaggact acttcaagaa
     1381 aatcgagtgc ttcgactccg tggaaatctc cggcgtggaa gatcggttca acgcctccct
     1441 gggcacatac cacgatctgc tgaaaattat caaggacaag gacttcctgg acaatgagga
     1501 aaacgaggac attctggaag atatcgtgct gaccctgaca ctgtttgagg acagagagat
     1561 gatcgaggaa cggctgaaaa cctatgccca cctgttcgac gacaaagtga tgaagcagct
     1621 gaagcggcgg agatacaccg gctggggcag gctgagccgg aagctgatca acggcatccg
     1681 ggacaagcag tccggcaaga caatcctgga tttcctgaag tccgacggct tcgccaacag
     1741 aaacttcatg cagctgatcc acgacgacag cctgaccttt aaagaggaca tccagaaagc
     1801 ccaggtgtcc ggccagggcg atagcctgca cgagcacatt gccaatctgg ccggcagccc
     1861 cgccattaag aagggcatcc tgcagacagt gaaggtggtg gacgagctcg tgaaagtgat
     1921 gggccggcac aagcccgaga acatcgtgat cgaaatggcc agagagaacc agaccaccca
     1981 gaagggacag aagaacagcc gcgagagaat gaagcggatc gaagagggca tcaaagagct
     2041 gggcagccag atcctgaaag aacaccccgt ggaaaacacc cagctgcaga acgagaagct
     2101 gtacctgtac tacctgcaga atgggcggga tatgtacgtg gaccaggaac tggacatcaa
     2161 ccggctgtcc gactacgatg tggaccatat cgtgcctcag agctttctga aggacgactc
     2221 catcgacaac aaggtgctga ccagaagcga caagaaccgg ggcaagagcg acaacgtgcc
     2281 ctccgaagag gtcgtgaaga agatgaagaa ctactggcgg cagctgctga acgccaagct
     2341 gattacccag agaaagttcg acaatctgac caaggccgag agaggcggcc tgagcgaact
     2401 ggataaggcc ggcttcatca agagacagct ggtggaaacc cggcagatca caaagcacgt
     2461 ggcacagatc ctggactccc ggatgaacac taagtacgac gagaatgaca agctgatccg
     2521 ggaagtgaaa gtgatcaccc tgaagtccaa gctggtgtcc gatttccgga aggatttcca
     2581 gttttacaaa gtgcgcgaga tcaacaacta ccaccacgcc cacgacgcct acctgaacgc
     2641 cgtcgtggga accgccctga tcaaaaagta ccctaagctg gaaagcgagt tcgtgtacgg
     2701 cgactacaag gtgtacgacg tgcggaagat gatcgccaag agcgagcagg aaatcggcaa
     2761 ggctaccgcc aagtacttct tctacagcaa catcatgaac tttttcaaga ccgagattac
     2821 cctggccaac ggcgagatcc ggaagcggcc tctgatcgag acaaacggcg aaaccgggga
     2881 gatcgtgtgg gataagggcc gggattttgc caccgtgcgg aaagtgctga gcatgcccca
     2941 agtgaatatc gtgaaaaaga ccgaggtgca gacaggcggc ttcagcaaag agtctatcct
     3001 gcccaagagg aacagcgata agctgatcgc cagaaagaag gactgggacc ctaagaagta
     3061 cggcggcttc gacagcccca ccgtggccta ttctgtgctg gtggtggcca aagtggaaaa
     3121 gggcaagtcc aagaaactga agagtgtgaa agagctgctg gggatcacca tcatggaaag
     3181 aagcagcttc gagaagaatc ccatcgactt tctggaagcc aagggctaca aagaagtgaa
     3241 aaaggacctg atcatcaagc tgcctaagta ctccctgttc gagctggaaa acggccggaa
     3301 gagaatgctg gcctctgccg gcgaactgca gaagggaaac gaactggccc tgccctccaa
     3361 atatgtgaac ttcctgtacc tggccagcca ctatgagaag ctgaagggct cccccgagga
     3421 taatgagcag aaacagctgt ttgtggaaca gcacaagcac tacctggacg agatcatcga
     3481 gcagatcagc gagttctcca agagagtgat cctggccgac gctaatctgg acaaagtgct
     3541 gtccgcctac aacaagcacc gggataagcc catcagagag caggccgaga atatcatcca
     3601 cctgtttacc ctgaccaatc tgggagcccc tgccgccttc aagtactttg acaccaccat
     3661 cgaccggaag aggtacacca gcaccaaaga ggtgctggac gccaccctga tccaccagag
     3721 catcaccggc ctgtacgaga cacggatcga cctgtctcag ctgggaggcg acaaaaggcc
     3781 ggcggccacg aaaaaggccg gccaggcaaa aaagaaaaag cttgagggca gaggaagtct
     3841 gctaacatgc ggtgacgtgg aggagaatcc cggccctgct agcatggtga gcaagggcga
     3901 ggaggataac atggccatca tcaaggagtt catgcgcttc aaggtgcaca tggagggctc
     3961 cgtgaacggc cacgagttcg agatcgaggg cgagggcgag ggccgcccct acgagggcac
     4021 ccagaccgcc aagctgaagg tgaccaaggg tggccccctg cccttcgcct gggacatcct
     4081 gtcccctcag ttcatgtacg gctccaaggc ctacgtgaag caccccgccg acatccccga
     4141 ctacttgaag ctgtccttcc ccgagggctt caagtgggag cgcgtgatga acttcgagga
     4201 cggcggcgtg gtgaccgtga cccaggactc ctccctgcag gacggcgagt tcatctacaa
     4261 ggtgaagctg cgcggcacca acttcccctc agacggcccc gtaatgcaga agaaaaccat
     4321 gggctgggag gcctcctccg agcggatgta ccccgaggac ggcgccctga agggcgagat
     4381 caagcagagg ctgaagctga aggacggcgg ccactacgac gctgaggtca agaccaccta
     4441 caaggccaag aagcccgtgc agctgcccgg cgcctacaac gtcaacatca agttggacat
     4501 cacctcccac aacgaggact acaccatcgt ggaacagtac gaacgcgccg agggccgcca
     4561 ctccaccggc ggcatggacg agctgtacaa gtaagaattc ctagagctcg ctgatcagcc
     4621 tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt gccttccttg
     4681 accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat tgcatcgcat
     4741 tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag caagggggag
     4801 gattgggaag agaatagcag gcatgctggg gagcggccgc aggaacccct agtgatggag
     4861 ttggccactc cctctctgcg cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc
     4921 cgacgcccgg gctttgcccg ggcggcctca gtgagcgagc gagcgcgcag ctgcctgcag
     4981 gggcgcctga tgcggtattt tctccttacg catctgtgcg gtatttcaca ccgcatacgt
     5041 caaagcaacc atagtacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     5101 cgcgcagcgt gaccgctaca cttgccagcg ccttagcgcc cgctcctttc gctttcttcc
     5161 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     5221 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg
     5281 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     5341 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaactct atctcgggct
     5401 attcttttga tttataaggg attttgccga tttcggtcta ttggttaaaa aatgagctga
     5461 tttaacaaaa atttaacgcg aattttaaca aaatattaac gtttacaatt ttatggtgca
     5521 ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac
     5581 ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga
     5641 ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac
     5701 gaaagggcct cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt
     5761 agacgtcagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     5821 aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataat
     5881 attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg
     5941 cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg
     6001 aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc
     6061 ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat
     6121 gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc cgcatacact
     6181 attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca
     6241 tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact
     6301 tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg
     6361 atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg
     6421 agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg
     6481 aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg
     6541 caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag
     6601 ccggtgagcg tggaagccgc ggtatcattg cagcactggg gccagatggt aagccctccc
     6661 gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga
     6721 tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat
     6781 atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc
     6841 tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag
     6901 accccgtaga aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct
     6961 gcttgcaaac aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac
     7021 caactctttt tccgaaggta actggcttca gcagagcgca gataccaaat actgttcttc
     7081 tagtgtagcc gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg
     7141 ctctgctaat cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt
     7201 tggactcaag acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt
     7261 gcacacagcc cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc
     7321 tatgagaaag cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca
     7381 gggtcggaac aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata
     7441 gtcctgtcgg gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg
     7501 ggcggagcct atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct
     7561 ggccttttgc tcacatgtga gggcctattt cccatgattc cttcatattt gcatatacga
     7621 tacaaggctg ttagagagat aattggaatt aatttgactg taaacacaaa gatattagta
     7681 caaaatacgt gacgtagaaa gtaataattt cttgggtagt ttgcagtttt aaaattatgt
     7741 tttaaaatgg actatcatat gcttaccgta acttgaaagt atttcgattt cttggcttta
     7801 tatatcttgt ggaaaggacg aaacaccgag atcgagtgcc gcatcacgtt ttagagctag
     7861 aaatagcaag ttaaaataag gctagtccgt tatcaacttg aaaaagtggc accgagtcgg
     7921 tgcttttttg ttttagagct agaaatagca agttaaaata aggctagtcc gtttttagcg
     7981 cgtgcgccaa ttctgcagac aaatggctct agaggtaccc gttacataac ttacggtaaa
     8041 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaatag taacgccaat
     8101 agggactttc cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt
     8161 acatcaagtg tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc
     8221 cgcctggcat tgtgcccagt acatgacctt atgggacttt cctacttggc agtacatcta
     8281 cgtattagtc atcgctatta ccatggtcga ggtgagcccc acgttctgct tcactctccc
     8341 catctccccc ccctccccac ccccaatttt gtatttattt attttttaat tattttgtgc
     8401 agcgatgggg gcgggggggg ggggggggcg cgcgccrggg gggggggggg ggggsggggg
     8461 gsggggsggg gsgaggcgga gaggtgcggc ggcagccaat cagagcggcg cgctccgaaa
     8521 gtttcctttt atggcgaggc ggcggcggcg gcggccctat aaaaagcgaa gcgcgcggcg
     8581 ggcgggagtc gctgcgcgct gccttcgccc cgtgccccgc tccgccgccg cctcgcgccg
     8641 cccgccccgg ctctgactga ccgcgttact cccacaggtg agcgggcggg acggcccttc
     8701 tcctccgggc tgtaattagc tgagcaagag gtaagggttt aagggatggt tggttggtgg
     8761 ggtattaatg tttaattacc tggagcacct gcctgaaatc actttttttc aggttggacc
     8821 ggtgccacca tggactataa ggaccacgac ggagactaca aggatcatga tattgattac
     8881 aaagacgatg acgataagat ggccccaaag aagaagcgga aggtcggtat ccacggagtc
     8941 ccagcagccg acaagaagta cagcatcggc ctggacatcg gcaccaactc tgtgggctgg
     9001 gccgtgatca ccgacgagta caaggtgccc agcaagaaat tcaaggtgct gggcaacacc
     9061 gaccggcaca gcatcaagaa gaacctgatc ggagccctgc tgttcgacag cggcgaaaca
     9121 gccgaggcca cccggctgaa gagaaccgcc agaagaagat acaccagacg gaagaaccgg
     9181 atctgctatc tgcaagagat cttcagcaac gagatggcca aggtggacga cagcttcttc
     9241 cacagactgg aagagtcctt cctggtggaa gaggataa
//
