LOCUS       sequence-84808-e        9674 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes CAS9 nuclease (GFP fusion) under control of the Toxoplasma 
            gondii SAG1 promotor. Vector also provides U6 controlled expression 
            of a single guide RNA for disruption of genes by CRISPR..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9674)
  AUTHORS   Shen B, Brown KM, Lee TD, Sibley LD
  TITLE     Efficient Gene Disruption in Diverse Strains of Toxoplasma gondii 
            Using CRISPR/CAS9.
  JOURNAL   MBio. 2014 May 13;5(3). pii: e01114-14. doi: 10.1128/mBio.01114-14.
  PUBMED    24825012
REFERENCE   2  (bases 1 to 9674)
  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: "MBio."; 
            date: "2014-05-13"; volume: "5(3). pii: e01114-14. doi"; pages: " 
            10.1128/mBio.01114-14"
COMMENT     Sequence Label: pSAG1::CAS9-U6::sgUPRT
FEATURES             Location/Qualifiers
     source          1..9674
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_RNA        559..633
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     promoter        complement(658..676)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(697..713)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(697..713)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(710..732)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    721..737
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(745..751,752..769,770..775))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    790..811
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(928..945)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1099..1687)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1179..1198)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(1858..2649,2650..2718))
                     /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     2481..2500
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2719..2823)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(2849..3304)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2986..3007)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3198..3217
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3431..3453
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     3445..3462
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     3446..3462
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3472..3491
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        3472..3490
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     3515..3531
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     3516..3532
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             4358..8461
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             4358..8461
                     /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="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             8465..8491
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             8471..8497
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /note="This is a variant of the standard HA tag, which has 
                     sequence YPYDVPDYA. Some anti-HA antibodies may not 
                     recognize this variant."
                     /translation="YDVPDYASL"
     CDS             8510..8530
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8537..8557
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(8597..8599,8600..9313)
                     /codon_start=1
                     /product="enhanced GFP (EGFP) derivative that exhibits 
                     robust folding and enhanced photostability (Zhong et al., 
                     2019)"
                     /label=SiriusGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTISFKDDG
                     TYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKA
                     NFKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     9247..9268
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     complement(9635..9651)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9635..9651)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
ORIGIN
        1 caagtaagca gaagcacgct gtatttccgg gagggtgcga cgagacaaag tgcgcgagtt
       61 gaaatcgtcg tggggacgat tgcaccgcgg ccacatgttg gagacactga ggacacacgg
      121 gaaacgcgaa agatttcaaa ttaacgtacc caagcgcgaa agcttgcgca gcatacactc
      181 gaagcgaaca tcccgaacca tcgagaggcg gagagcgata agtctttcac gctgcgaagt
      241 gttgcgacgg ctgcgccgct gcactgtgaa ttgggcgcca atattgcatc ctaggcctga
      301 cgcgcctcct gcagaacgcg agacactggg atatgtagag ccaaggggga aaccttcgaa
      361 ctctcgaatg tcttctctga caagaatcat atttccatca gttctgtcag attttcaaat
      421 ggcgacctgc agaggcctgc ttcctccctg tgcgctcttc gaaggggctt tctgtcgcgc
      481 agggtcacct cgtccccgaa gggggtgttt gccttctggt aaatggggat gtcaagttgg
      541 cgtctcgatt gtgagagcgt tttagagcta gaaatagcaa gttaaaataa ggctagtccg
      601 ttatcaactt gaaaagtggc accgagtcgg tgcttttttg agctccagct tttgttccct
      661 ttagtgaggg ttaatttcga gcttggcgta atcatggtca tagctgtttc ctgtgtgaaa
      721 ttgttatccg ctcacaattc cacacaacat acgagccgga agcataaagt gtaaagcctg
      781 gggtgcctaa tgagtgagct aactcacatt aattgcgttg cgctcactgc ccgctttcca
      841 gtcgggaaac ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg
      901 tttgcgtatt gggcgctctt ccgcttcctc gctcactgac tcgctgcgct cggtcgttcg
      961 gctgcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
     1021 ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa
     1081 ggccgcgttg ctggcgtttt tccataggct ccgcccccct gacgagcatc acaaaaatcg
     1141 acgctcaagt cagaggtggc gaaacccgac aggactataa agataccagg cgtttccccc
     1201 tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg cttaccggat acctgtccgc
     1261 ctttctccct tcgggaagcg tggcgctttc tcatagctca cgctgtaggt atctcagttc
     1321 ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa ccccccgttc agcccgaccg
     1381 ctgcgcctta tccggtaact atcgtcttga gtccaacccg gtaagacacg acttatcgcc
     1441 actggcagca gccactggta acaggattag cagagcgagg tatgtaggcg gtgctacaga
     1501 gttcttgaag tggtggccta actacggcta cactagaagg acagtatttg gtatctgcgc
     1561 tctgctgaag ccagttacct tcggaaaaag agttggtagc tcttgatccg gcaaacaaac
     1621 caccgctggt agcggtggtt tttttgtttg caagcagcag attacgcgca gaaaaaaagg
     1681 atctcaagaa gatcctttga tcttttctac ggggtctgac gctcagtgga acgaaaactc
     1741 acgttaaggg attttggtca tgagattatc aaaaaggatc ttcacctaga tccttttaaa
     1801 ttaaaaatga agttttaaat caatctaaag tatatatgag taaacttggt ctgacagtta
     1861 ccaatgctta atcagtgagg cacctatctc agcgatctgt ctatttcgtt catccatagt
     1921 tgcctgactc cccgtcgtgt agataactac gatacgggag ggcttaccat ctggccccag
     1981 tgctgcaatg ataccgcgag acccacgctc accggctcca gatttatcag caataaacca
     2041 gccagccgga agggccgagc gcagaagtgg tcctgcaact ttatccgcct ccatccagtc
     2101 tattaattgt tgccgggaag ctagagtaag tagttcgcca gttaatagtt tgcgcaacgt
     2161 tgttgccatt gctacaggca tcgtggtgtc acgctcgtcg tttggtatgg cttcattcag
     2221 ctccggttcc caacgatcaa ggcgagttac atgatccccc atgttgtgca aaaaagcggt
     2281 tagctccttc ggtcctccga tcgttgtcag aagtaagttg gccgcagtgt tatcactcat
     2341 ggttatggca gcactgcata attctcttac tgtcatgcca tccgtaagat gcttttctgt
     2401 gactggtgag tactcaacca agtcattctg agaatagtgt atgcggcgac cgagttgctc
     2461 ttgcccggcg tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat
     2521 cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag
     2581 ttcgatgtaa cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt
     2641 ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg
     2701 gaaatgttga atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta
     2761 ttgtctcatg agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc
     2821 gcgcacattt ccccgaaaag tgccacctaa attgtaagcg ttaatatttt gttaaaattc
     2881 gcgttaaatt tttgttaaat cagctcattt tttaaccaat aggccgaaat cggcaaaatc
     2941 ccttataaat caaaagaata gaccgagata gggttgagtg ttgttccagt ttggaacaag
     3001 agtccactat taaagaacgt ggactccaac gtcaaagggc gaaaaaccgt ctatcagggc
     3061 gatggcccac tacgtgaacc atcaccctaa tcaagttttt tggggtcgag gtgccgtaaa
     3121 gcactaaatc ggaaccctaa agggagcccc cgatttagag cttgacgggg aaagccggcg
     3181 aacgtggcga gaaaggaagg gaagaaagcg aaaggagcgg gcgctagggc gctggcaagt
     3241 gtagcggtca cgctgcgcgt aaccaccaca cccgccgcgc ttaatgcgcc gctacagggc
     3301 gcgtcccatt cgccattcag gctgcgcaac tgttgggaag ggcgatcggt gcgggcctct
     3361 tcgctattac gccagctggc gaaaggggga tgtgctgcaa ggcgattaag ttgggtaacg
     3421 ccagggtttt cccagtcacg acgttgtaaa acgacggcca gtgagcgcgc gtaatacgac
     3481 tcactatagg gcgaattggg taccgggccc cccctcgagg tcgacggtat cgataagctt
     3541 ttacatccgt tgccttttcc acggtccgtg atttcatgtg cgtgcagctt caaagactgg
     3601 tcgttgcgac taataagact gcagtgacag gtcgaatggt gggcaccttg ctgatgacta
     3661 tctactgcaa agtctgagac aacgaacgaa acttcccaca cgaggcattt gaaactgacg
     3721 gtgtctaggt aatatgcact gcaagacacg gtactggggc ctcgctgaat taggggccga
     3781 tctcgttgcc ctatcagtgc tcacagtgcc gcaacgtaac accagggcag gttcttgaca
     3841 gtggcaacaa tgtgcgacgg gcgtgtgaac gtttcgtagt catagcgcta gcacgtacct
     3901 agccacatgg tcgtgaggag ctttaccatg cgtctagaag gtggatgcgg gacacgcctt
     3961 cctggccttt ggctcccgag acgcgtgttc taaccacaaa ccttgagacg cgtgttccaa
     4021 ccacgcaccc tgacacgcgt gttccaacca cgcaccctga gacgcgtgtt ctaaccacgc
     4081 accctgagac gcgtgttcta accacgcacc ctgagacgcg tgttctgccg cacaatgtgc
     4141 acctgtagga agctgtagtc actgctgatt ctcactgttc tcggcaaggg ccgacgaccg
     4201 gagtacagtt tttgtgggca gagccgttgt gcagctttcc gttcttctcg gttgtgtcac
     4261 atgtgtcatt gtcgtgtaaa cacacggttg tatgtcggtt tcgctgcacc acttcattat
     4321 ttcttctggt tttttgacga gtatgcatct agacaaaatg gacaagaagt acagcatcgg
     4381 cctggacatc ggcaccaact ctgtgggctg ggccgtgatc accgacgagt acaaggtgcc
     4441 cagcaagaaa ttcaaggtgc tgggcaacac cgaccggcac agcatcaaga agaacctgat
     4501 cggcgccctg ctgttcgaca gcggagaaac agccgaggcc acccggctga agagaaccgc
     4561 cagaagaaga tacaccagac ggaagaaccg gatctgctat ctgcaagaga tcttcagcaa
     4621 cgagatggcc aaggtggacg acagcttctt ccacagactg gaagagtcct tcctggtgga
     4681 agaggataag aagcacgagc ggcaccccat cttcggcaac atcgtggacg aggtggccta
     4741 ccacgagaag taccccacca tctaccacct gagaaagaaa ctggtggaca gcaccgacaa
     4801 ggccgacctg cggctgatct atctggccct ggcccacatg atcaagttcc ggggccactt
     4861 cctgatcgag ggcgacctga accccgacaa cagcgacgtg gacaagctgt tcatccagct
     4921 ggtgcagacc tacaaccagc tgttcgagga aaaccccatc aacgccagcg gcgtggacgc
     4981 caaggccatc ctgtctgcca gactgagcaa gagcagacgg ctggaaaatc tgatcgccca
     5041 gctgcccggc gagaagaaga atggcctgtt cggcaacctg attgccctga gcctgggcct
     5101 gacccccaac ttcaagagca acttcgacct ggccgaggat gccaaactgc agctgagcaa
     5161 ggacacctac gacgacgacc tggacaacct gctggcccag atcggcgacc agtacgccga
     5221 cctgtttctg gccgccaaga acctgtccga cgccatcctg ctgagcgaca tcctgagagt
     5281 gaacaccgag atcaccaagg cccccctgag cgcctctatg atcaagagat acgacgagca
     5341 ccaccaggac ctgaccctgc tgaaagctct cgtgcggcag cagctgcctg agaagtacaa
     5401 agagattttc ttcgaccaga gcaagaacgg ctacgccggc tacatcgatg gcggagccag
     5461 ccaggaagag ttctacaagt tcatcaagcc catcctggaa aagatggacg gcaccgagga
     5521 actgctcgtg aagctgaaca gagaggacct gctgcggaag cagcggacct tcgacaacgg
     5581 cagcatcccc caccagatcc acctgggaga gctgcacgcc attctgcggc ggcaggaaga
     5641 tttttaccca ttcctgaagg acaaccggga aaagatcgag aagatcctga ccttccgcat
     5701 cccctactac gtgggccctc tggccagggg aaacagcaga ttcgcctgga tgaccagaaa
     5761 gagcgaggaa accatcaccc cctggaactt cgaggaagtg gtggacaagg gcgccagcgc
     5821 ccagagcttc atcgagcgga tgaccaactt cgataagaac ctgcccaacg agaaggtgct
     5881 gcccaagcac agcctgctgt acgagtactt caccgtgtac aacgagctga ccaaagtgaa
     5941 atacgtgacc gagggaatga gaaagcccgc cttcctgagc ggcgagcaga aaaaagccat
     6001 cgtggacctg ctgttcaaga ccaaccggaa agtgaccgtg aagcagctga aagaggacta
     6061 cttcaagaaa atcgagtgct tcgactccgt ggaaatctcc ggcgtggaag atcggttcaa
     6121 cgcctccctg ggcacatacc acgatctgct gaaaattatc aaggacaagg acttcctgga
     6181 caatgaggaa aacgaggaca ttctggaaga tatcgtgctg accctgacac tgtttgagga
     6241 cagagagatg atcgaggaac ggctgaaaac ctatgcccac ctgttcgacg acaaagtgat
     6301 gaagcagctg aagcggcgga gatacaccgg ctggggcagg ctgagccgga agctgatcaa
     6361 cggcatccgg gacaagcagt ccggcaagac aatcctggat ttcctgaagt ccgacggctt
     6421 cgccaacaga aacttcatgc agctgatcca cgacgacagc ctgaccttta aagaggacat
     6481 ccagaaagcc caggtgtccg gccagggcga tagcctgcac gagcacattg ccaatctggc
     6541 cggcagcccc gccattaaga agggcatcct gcagacagtg aaggtggtgg acgagctcgt
     6601 gaaagtgatg ggccggcaca agcccgagaa catcgtgatc gaaatggcca gagagaacca
     6661 gaccacccag aagggacaga agaacagccg cgagagaatg aagcggatcg aagagggcat
     6721 caaagagctg ggcagccaga tcctgaaaga acaccccgtg gaaaacaccc agctgcagaa
     6781 cgagaagctg tacctgtact acctgcagaa tgggcgggat atgtacgtgg accaggaact
     6841 ggacatcaac cggctgtccg actacgatgt ggaccatatc gtgcctcaga gctttctgaa
     6901 ggacgactcc atcgataaca aagtgctgac tcggagcgac aagaaccggg gcaagagcga
     6961 caacgtgccc tccgaagagg tcgtgaagaa gatgaagaac tactggcgcc agctgctgaa
     7021 tgccaagctg attacccaga ggaagttcga caatctgacc aaggccgaga gaggcggcct
     7081 gagcgaactg gataaggccg gcttcatcaa gagacagctg gtggaaaccc ggcagatcac
     7141 aaagcacgtg gcacagatcc tggactcccg gatgaacact aagtacgacg agaacgacaa
     7201 actgatccgg gaagtgaaag tgatcaccct gaagtccaag ctggtgtccg atttccggaa
     7261 ggatttccag ttttacaaag tgcgcgagat caacaactac caccacgccc acgacgccta
     7321 cctgaacgcc gtcgtgggaa ccgccctgat caaaaagtac cctaagctgg aaagcgagtt
     7381 cgtgtacggc gactacaagg tgtacgacgt gcggaagatg atcgccaaga gcgagcagga
     7441 aatcggcaag gctaccgcca agtacttctt ctacagcaac atcatgaact ttttcaagac
     7501 cgagattacc ctggccaacg gcgagatccg gaagcggcct ctgatcgaga caaacggcga
     7561 aacaggcgag atcgtgtggg ataagggccg ggactttgcc accgtgcgga aagtgctgtc
     7621 tatgccccaa gtgaatatcg tgaaaaagac cgaggtgcag acaggcggct tcagcaaaga
     7681 gtctatcctg cccaagagga acagcgacaa gctgatcgcc agaaagaagg actgggaccc
     7741 taagaagtac ggcggcttcg acagccccac cgtggcctat tctgtgctgg tggtggccaa
     7801 agtggaaaag ggcaagtcca agaaactgaa gagtgtgaaa gagctgctgg ggatcaccat
     7861 catggaaaga agcagcttcg agaagaatcc catcgacttt ctggaagcca agggctacaa
     7921 agaagtgaaa aaggacctga tcatcaagct gcctaagtac tccctgttcg agctggaaaa
     7981 cggccggaag agaatgctgg cctctgccgg cgaactgcag aagggaaacg aactggccct
     8041 gccctccaaa tatgtgaact tcctgtacct ggccagccac tatgagaagc tgaagggctc
     8101 ccccgaggat aatgagcaga aacagctgtt tgtggaacag cacaaacact acctggacga
     8161 gatcatcgag cagatcagcg agttctccaa gagagtgatc ctggccgacg ctaatctgga
     8221 caaggtgctg agcgcctaca acaagcacag agacaagcct atcagagagc aggccgagaa
     8281 tatcatccac ctgtttaccc tgaccaatct gggagcccct gccgccttca agtactttga
     8341 caccaccatc gaccggaaga ggtacaccag caccaaagag gtgctggacg ccaccctgat
     8401 ccaccagagc atcaccggcc tgtacgagac acggatcgac ctgtctcagc tgggaggcga
     8461 cgcctatccc tatgacgtgc ccgattatgc cagcctgggc agcggctccc ccaagaaaaa
     8521 acgcaaggtg gaagatccta agaaaaagcg gaaagtggac ggcattggta gtgggagcaa
     8581 cggcagcagc ggatccgtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     8641 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgcgcggcg agggcgaggg
     8701 cgatgccacc aacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     8761 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     8821 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     8881 gcgcaccatc tccttcaagg acgacggcac ctacaagacc cgcgccgagg tgaagttcga
     8941 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     9001 catcctgggg cacaagctgg agtacaactt caacagccac aacgtctata tcacggccga
     9061 caagcagaag aacggcatca aggcgaactt caagatccgc cacaacgtcg aggacggcag
     9121 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     9181 gcccgacaac cactacctga gcacccagtc caagctgagc aaagacccca acgagaagcg
     9241 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     9301 gctgtacaag tagttaatta atcaccgttg tgctcacttc tcaaatcgac aaaggaaaca
     9361 cacttcgtgc agcatgtgcc ccattataaa gaaactgagt tgttccgttg tggcttgcag
     9421 gtgtcacatc cacaaaaacc ggccgactct aaataggagt gtttcgcagc aagcagcgaa
     9481 agtttatgac tgggtccgaa tctctgaacg gatgtgtggc ggacctggct gatgttgatc
     9541 gccgtcgaca cacgcgccac atgggtcaat acacaagaca gctatcagtt gttttagtcg
     9601 aaccggttaa cacaattctt gcccccccga gggggatcca ctagttctag agcggccgcc
     9661 accgcggtgg agct
//
