LOCUS       sequence-166589-       10347 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses S. pyogenes FokI-dCas9-NLS with a 25 amino acid linker 
            (GGGGS)5 fusion. Also expresses Csy4 for cleavage of multiplexed 
            gRNA transcripts. Derived from pSQT1601 (Addgene #53369)..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10347)
  AUTHORS   Havlicek S, Shen Y, Alpagu Y, Bruntraeger MB, Zufir NB, Phuah ZY, Fu
            Z, Dunn NR, Stanton LW
  TITLE     Re-engineered RNA-Guided FokI-Nucleases for Improved Genome Editing 
            in Human Cells.
  JOURNAL   Mol Ther. 2017 Feb 1;25(2):342-355. doi: 
            10.1016/j.ymthe.2016.11.007.
  PUBMED    28153087
REFERENCE   2  (bases 1 to 10347)
  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: 
            "10.1016/j.ymthe.2016.11"; journalName: "Mol Ther"; date: 
            "2017-02-1- 1"; volume: "25"; issue: "2"; pages: "342-355"
COMMENT     Sequence Label: pSH-Csy4-T2A-SpRFN
FEATURES             Location/Qualifiers
     source          1..10347
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        4..383
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        386..661
                     /label=chicken beta-actin promoter
     intron          662..1670
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1594..1616
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1678..1697
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     regulatory      1763..1772
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2333..2386
                     /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             2390..2410
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2417..3004
                     /codon_start=1
                     /product="nonspecific DNA cleavage domain of the FokI 
                     endonuclease (Li et al., 1992)"
                     /label=FokI cleavage domain
                     /note="must dimerize to be catalytically active"
                     /translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV
                     MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQR
                     YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
                     SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
     CDS             join(3080..6478,6479..6481,6482..7078,7079,7080..7082,
                     7083..7084,7085..7087,7088..7180)
                     /codon_start=1
                     /product="Streptococcus pyogenes Cas9 endonuclease with the
                     D1135V/R1335Q/T1337R mutations"
                     /label=Cas9 VQR
                     /note="has an altered recognition specificity for the 
                     protospacer adjacent motif (PAM) (Kleinstiver et al., 
                     2015)"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             join(3080..3103,3104..3106,3107..5593,5594..5596,
                     5597..7180)
                     /codon_start=1
                     /product="catalytically dead mutant of the Cas9 
                     endonuclease from the Streptococcus pyogenes Type II 
                     CRISPR/Cas system"
                     /label=dCas9
                     /note="RNA-guided DNA-binding protein that lacks 
                     endonuclease activity due to the D10A mutation in the RuvC 
                     catalytic domain and the H840A mutation in the HNH 
                     catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDAIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     CDS             7187..7207
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             7214..7279
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     primer_bind     complement(7369..7388)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    7434..7489
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(7488..7507)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(7850..7866)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(7850..7866)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(7863..7885)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    7874..7890
                     /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(7898..7904,7905..7922,7923..7928))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    7943..7964
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        8023..8218
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      8069..8204
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    8224..8358
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(8261..8280)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     8315..8334
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(8426..8443)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(8597..9185)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(8677..8696)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(9356..10147,10148..10216))
                     /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     9979..9998
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(10217..10321)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcgacattg attattgact agttattaat agtaatcaat tacggggtca ttagttcata
       61 gcccatatat ggagttccgc gttacataac ttacggtaaa tggcccgcct ggctgaccgc
      121 ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag
      181 ggactttcca ttgacgtcaa tgggtggact atttacggta aactgcccac ttggcagtac
      241 atcaagtgta tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg
      301 cctggcatta tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg
      361 tattagtcat cgctattacc atgggtcgag gtgagcccca cgttctgctt cactctcccc
      421 atctcccccc cctccccacc cccaattttg tatttattta ttttttaatt attttgtgca
      481 gcgatggggg cggggggggg gggggcgcgc gccaggcggg gcggggcggg gcgaggggcg
      541 gggcggggcg aggcggagag gtgcggcggc agccaatcag agcggcgcgc tccgaaagtt
      601 tccttttatg gcgaggcggc ggcggcggcg gccctataaa aagcgaagcg cgcggcgggc
      661 gggagtcgct gcgttgcctt cgccccgtgc cccgctccgc gccgcctcgc gccgcccgcc
      721 ccggctctga ctgaccgcgt tactcccaca ggtgagcggg cgggacggcc cttctcctcc
      781 gggctgtaat tagcgcttgg tttaatgacg gctcgtttct tttctgtggc tgcgtgaaag
      841 ccttaaaggg ctccgggagg gccctttgtg cgggggggag cggctcgggg ggtgcgtgcg
      901 tgtgtgtgtg cgtggggagc gccgcgtgcg gcccgcgctg cccggcggct gtgagcgctg
      961 cgggcgcggc gcggggcttt gtgcgctccg cgtgtgcgcg aggggagcgc ggccgggggc
     1021 ggtgccccgc ggtgcggggg ggctgcgagg ggaacaaagg ctgcgtgcgg ggtgtgtgcg
     1081 tgggggggtg agcagggggt gtgggcgcgg cggtcgggct gtaacccccc cctgcacccc
     1141 cctccccgag ttgctgagca cggcccggct tcgggtgcgg ggctccgtgc ggggcgtggc
     1201 gcggggctcg ccgtgccggg cggggggtgg cggcaggtgg gggtgccggg cggggcgggg
     1261 ccgcctcggg ccggggaggg ctcgggggag gggcgcggcg gccccggagc gccggcggct
     1321 gtcgaggcgc ggcgagccgc agccattgcc ttttatggta atcgtgcgag agggcgcagg
     1381 gacttccttt gtcccaaatc tggcggagcc gaaatctggg aggcgccgcc gcaccccctc
     1441 tagcgggcgc gggcgaagcg gtgcggcgcc ggcaggaagg aaatgggcgg ggagggcctt
     1501 cgtgcgtcgc cgcgccgccg tccccttctc catctccagc ctcggggctg ccgcaggggg
     1561 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1621 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1681 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attctgcagt cgacggtacc
     1741 gcgggcccgg gatccaccgg tcgccaccat gggtgatcat tatctggata ttcggctgag
     1801 gcctgatcca gagttcccac ctgcgcagct gatgtctgtc ctttttggca aacttcatca
     1861 ggccctggtt gcccagggcg gagatcggat aggggtaagc tttccagacc tcgacgaaag
     1921 ccggagccgc ctgggagaac gcctgcggat ccacgcttct gccgacgatc tgagagcctt
     1981 gctggcaagg ccatggcttg aggggctccg ggatcacctg cagtttggcg aacccgccgt
     2041 tgttccccac ccaacccctt atcggcaggt gtctagagtg caggccaaat ctaatccaga
     2101 acggctgcga cggcgactca tgcggcgaca tgatcttagc gaggaagagg cccgaaaaag
     2161 aatccctgat accgtggccc gcgcccttga cttgcctttt gtcacactgc ggtcccagag
     2221 tacggggcag catttcagac ttttcattcg acacgggcca ctgcaagtta ccgccgaaga
     2281 aggaggcttt acttgttatg gactctccaa gggaggtttc gtgccctggt ttgagggcag
     2341 aggaagtctg ttaacatgcg gtgacgtcga ggagaatcct ggcccaatgc ctaagaagaa
     2401 gcggaaggtg agcagccaac ttgtgaagtc tgaactcgag gagaaaaaat cagagttgag
     2461 acacaagttg aagtacgtgc cacacgaata catcgagctt atcgagatcg ccagaaacag
     2521 tacccaggat aggatccttg agatgaaagt catggagttc tttatgaagg tctacggtta
     2581 tagaggaaag caccttggcg gtagcagaaa gcccgatggc gccatctata ctgtcggatc
     2641 tcctatcgat tatggggtga tcgtggatac caaagcttac tcaggcgggt acaacttgcc
     2701 cataggacaa gccgacgaga tgcagcggta tgtcgaagag aaccagacgc gcaacaagca
     2761 catcaacccc aatgaatggt ggaaagtgta cccaagtagt gtgactgagt tcaagttcct
     2821 gtttgtctcc ggccacttta agggcaatta taaagctcag ctcactagac tcaatcacat
     2881 cacaaactgc aacggagctg tgttgtcagt ggaggagctc ctgattggag gcgagatgat
     2941 caaagccggc acccttacac tggaggaggt gcggcggaag ttcaacaatg gagagatcaa
     3001 cttcggtggc ggtggatccg gtggcggtgg atccggtggc ggtggatccg gtggcggtgg
     3061 atccggtggt ggtggatccg ataaaaagta ttctattggt ttagccatcg gcactaattc
     3121 cgttggatgg gctgtcataa ccgatgaata caaagtacct tcaaagaaat ttaaggtgtt
     3181 ggggaacaca gaccgtcatt cgattaaaaa gaatcttatc ggtgccctcc tattcgatag
     3241 tggcgaaacg gcagaggcga ctcgcctgaa acgaaccgct cggagaaggt atacacgtcg
     3301 caagaaccga atatgttact tacaagaaat ttttagcaat gagatggcca aagttgacga
     3361 ttctttcttt caccgtttgg aagagtcctt ccttgtcgaa gaggacaaga aacatgaacg
     3421 gcaccccatc tttggaaaca tagtagatga ggtggcatat catgaaaagt acccaacgat
     3481 ttatcacctc agaaaaaagc tagttgactc aactgataaa gcggacctga ggttaatcta
     3541 cttggctctt gcccatatga taaagttccg tgggcacttt ctcattgagg gtgatctaaa
     3601 tccggacaac tcggatgtcg acaaactgtt catccagtta gtacaaacct ataatcagtt
     3661 gtttgaagag aaccctataa atgcaagtgg cgtggatgcg aaggctattc ttagcgcccg
     3721 cctctctaaa tcccgacggc tagaaaacct gatcgcacaa ttacccggag agaagaaaaa
     3781 tgggttgttc ggtaacctta tagcgctctc actaggcctg acaccaaatt ttaagtcgaa
     3841 cttcgactta gctgaagatg ccaaattgca gcttagtaag gacacgtacg atgacgatct
     3901 cgacaatcta ctggcacaaa ttggagatca gtatgcggac ttatttttgg ctgccaaaaa
     3961 ccttagcgat gcaatcctcc tatctgacat actgagagtt aatactgaga ttaccaaggc
     4021 gccgttatcc gcttcaatga tcaaaaggta cgatgaacat caccaagact tgacacttct
     4081 caaggcccta gtccgtcagc aactgcctga gaaatataag gaaatattct ttgatcagtc
     4141 gaaaaacggg tacgcaggtt atattgacgg cggagcgagt caagaggaat tctacaagtt
     4201 tatcaaaccc atattagaga agatggatgg gacggaagag ttgcttgtaa aactcaatcg
     4261 cgaagatcta ctgcgaaagc agcggacttt cgacaacggt agcattccac atcaaatcca
     4321 cttaggcgaa ttgcatgcta tacttagaag gcaggaggat ttttatccgt tcctcaaaga
     4381 caatcgtgaa aagattgaga aaatcctaac ctttcgcata ccttactatg tgggacccct
     4441 ggcccgaggg aactctcggt tcgcatggat gacaagaaag tccgaagaaa cgattactcc
     4501 atggaatttt gaggaagttg tcgataaagg tgcgtcagct caatcgttca tcgagaggat
     4561 gaccaacttt gacaagaatt taccgaacga aaaagtattg cctaagcaca gtttacttta
     4621 cgagtatttc acagtgtaca atgaactcac gaaagttaag tatgtcactg agggcatgcg
     4681 taaacccgcc tttctaagcg gagaacagaa gaaagcaata gtagatctgt tattcaagac
     4741 caaccgcaaa gtgacagtta agcaattgaa agaggactac tttaagaaaa ttgaatgctt
     4801 cgattctgtc gagatctccg gggtagaaga tcgatttaat gcgtcacttg gtacgtatca
     4861 tgacctccta aagataatta aagataagga cttcctggat aacgaagaga atgaagatat
     4921 cttagaagat atagtgttga ctcttaccct ctttgaagat cgggaaatga ttgaggaaag
     4981 actaaaaaca tacgctcacc tgttcgacga taaggttatg aaacagttaa agaggcgtcg
     5041 ctatacgggc tggggacgat tgtcgcggaa acttatcaac gggataagag acaagcaaag
     5101 tggtaaaact attctcgatt ttctaaagag cgacggcttc gccaatagga actttatgca
     5161 gctgatccat gatgactctt taaccttcaa agaggatata caaaaggcac aggtttccgg
     5221 acaaggggac tcattgcacg aacatattgc gaatcttgct ggttcgccag ccatcaaaaa
     5281 gggcatactc cagacagtca aagtagtgga tgagctagtt aaggtcatgg gacgtcacaa
     5341 accggaaaac attgtaatcg agatggcacg cgaaaatcaa acgactcaga aggggcaaaa
     5401 aaacagtcga gagcggatga agagaataga agagggtatt aaagaactgg gcagccagat
     5461 cttaaaggag catcctgtgg aaaataccca attgcagaac gagaaacttt acctctatta
     5521 cctacaaaat ggaagggaca tgtatgttga tcaggaactg gacataaacc gtttatctga
     5581 ttacgacgtc gatgccattg taccccaatc ctttttgaag gacgattcaa tcgacaataa
     5641 agtgcttaca cgctcggata agaaccgagg gaaaagtgac aatgttccaa gcgaggaagt
     5701 cgtaaagaaa atgaagaact attggcggca gctcctaaat gcgaaactga taacgcaaag
     5761 aaagttcgat aacttaacta aagctgagag gggtggcttg tctgaacttg acaaggccgg
     5821 atttattaaa cgtcagctcg tggaaacccg ccaaatcaca aagcatgttg cacagatact
     5881 agattcccga atgaatacga aatacgacga gaacgataag ctgattcggg aagtcaaagt
     5941 aatcacttta aagtcaaaat tggtgtcgga cttcagaaag gattttcaat tctataaagt
     6001 tagggagata aataactacc accatgcgca cgacgcttat cttaatgccg tcgtagggac
     6061 cgcactcatt aagaaatacc cgaagctaga aagtgagttt gtgtatggtg attacaaagt
     6121 ttatgacgtc cgtaagatga tcgcgaaaag cgaacaggag ataggcaagg ctacagccaa
     6181 atacttcttt tattctaaca ttatgaattt ctttaagacg gaaatcactc tggcaaacgg
     6241 agagatacgc aaacgacctt taattgaaac caatggggag acaggtgaaa tcgtatggga
     6301 taagggccgg gacttcgcga cggtgagaaa agttttgtcc atgccccaag tcaacatagt
     6361 aaagaaaact gaggtgcaga ccggagggtt ttcaaaggaa tcgattcttc caaaaaggaa
     6421 tagtgataag ctcatcgctc gtaaaaagga ctgggacccg aaaaagtacg gtggcttcga
     6481 tagccctaca gttgcctatt ctgtcctagt agtggcaaaa gttgagaagg gaaaatccaa
     6541 gaaactgaag tcagtcaaag aattattggg gataacgatt atggagcgct cgtcttttga
     6601 aaagaacccc atcgacttcc ttgaggcgaa aggttacaag gaagtaaaaa aggatctcat
     6661 aattaaacta ccaaagtata gtctgtttga gttagaaaat ggccgaaaac ggatgttggc
     6721 tagcgccgga gagcttcaaa aggggaacga actcgcacta ccgtctaaat acgtgaattt
     6781 cctgtattta gcgtcccatt acgagaagtt gaaaggttca cctgaagata acgaacagaa
     6841 gcaacttttt gttgagcagc acaaacatta tctcgacgaa atcatagagc aaatttcgga
     6901 attcagtaag agagtcatcc tagctgatgc caatctggac aaagtattaa gcgcatacaa
     6961 caagcacagg gataaaccca tacgtgagca ggcggaaaat attatccatt tgtttactct
     7021 taccaacctc ggcgctccag ccgcattcaa gtattttgac acaacgatag atcgcaaacg
     7081 atacacttct accaaggagg tgctagacgc gacactgatt caccaatcca tcacgggatt
     7141 atatgaaact cggatagatt tgtcacagct tgggggtgac ggatccccca agaagaagag
     7201 gaaagtctcg agcgactaca aagaccatga cggtgattat aaagatcatg acatcgatta
     7261 caaggatgac gatgacaagt gaagcggccg cactcctcag gtgcaggctg cctatcagaa
     7321 ggtggtggct ggtgtggcca atgccctggc tcacaaatac cactgagatc tttttccctc
     7381 tgccaaaaat tatggggaca tcatgaagcc ccttgagcat ctgacttctg gctaataaag
     7441 gaaatttatt ttcattgcaa tagtgtgttg gaattttttg tgtctctcac tcggaaggac
     7501 atatgggagg gcaaatcatt taaaacatca gaatgagtat ttggtttaga gtttggcaac
     7561 atatgcccat atgctggctg ccatgaacaa aggttggcta taaagaggtc atcagtatat
     7621 gaaacagccc cctgctgtcc attccttatt ccatagaaaa gccttgactt gaggttagat
     7681 tttttttata ttttgttttg tgttattttt ttctttaaca tccctaaaat tttccttaca
     7741 tgttttacta gccagatttt tcctcctctc ctgactactc ccagtcatag ctgtccctct
     7801 tctcttatga agatccctcg acctgcagcc caagcttggc gtaatcatgg tcatagctgt
     7861 ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc ggaagcataa
     7921 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     7981 tgcccgcttt ccagtcggga aacctgtcgt gccagcggat ccgcatctca attagtcagc
     8041 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     8101 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctcggc
     8161 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     8221 gctaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     8281 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     8341 tcttatcatg tctggatccg ctgcattaat gaatcggcca acgcgcgggg agaggcggtt
     8401 tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc
     8461 tgcggcgagc ggtatcagct cactcaaagg cggtaatacg gttatccaca gaatcagggg
     8521 ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa ggccaggaac cgtaaaaagg
     8581 ccgcgttgct ggcgtttttc cataggctcc gcccccctga cgagcatcac aaaaatcgac
     8641 gctcaagtca gaggtggcga aacccgacag gactataaag ataccaggcg tttccccctg
     8701 gaagctccct cgtgcgctct cctgttccga ccctgccgct taccggatac ctgtccgcct
     8761 ttctcccttc gggaagcgtg gcgctttctc aatgctcacg ctgtaggtat ctcagttcgg
     8821 tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc ccccgttcag cccgaccgct
     8881 gcgccttatc cggtaactat cgtcttgagt ccaacccggt aagacacgac ttatcgccac
     8941 tggcagcagc cactggtaac aggattagca gagcgaggta tgtaggcggt gctacagagt
     9001 tcttgaagtg gtggcctaac tacggctaca ctagaaggac agtatttggt atctgcgctc
     9061 tgctgaagcc agttaccttc ggaaaaagag ttggtagctc ttgatccggc aaacaaacca
     9121 ccgctggtag cggtggtttt tttgtttgca agcagcagat tacgcgcaga aaaaaaggat
     9181 ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc tcagtggaac gaaaactcac
     9241 gttaagggat tttggtcatg agattatcaa aaaggatctt cacctagatc cttttaaatt
     9301 aaaaatgaag ttttaaatca atctaaagta tatatgagta aacttggtct gacagttacc
     9361 aatgcttaat cagtgaggca cctatctcag cgatctgtct atttcgttca tccatagttg
     9421 cctgactccc cgtcgtgtag ataactacga tacgggaggg cttaccatct ggccccagtg
     9481 ctgcaatgat accgcgagac ccacgctcac cggctccaga tttatcagca ataaaccagc
     9541 cagccggaag ggccgagcgc agaagtggtc ctgcaacttt atccgcctcc atccagtcta
     9601 ttaattgttg ccgggaagct agagtaagta gttcgccagt taatagtttg cgcaacgttg
     9661 ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt tggtatggct tcattcagct
     9721 ccggttccca acgatcaagg cgagttacat gatcccccat gttgtgcaaa aaagcggtta
     9781 gctccttcgg tcctccgatc gttgtcagaa gtaagttggc cgcagtgtta tcactcatgg
     9841 ttatggcagc actgcataat tctcttactg tcatgccatc cgtaagatgc ttttctgtga
     9901 ctggtgagta ctcaaccaag tcattctgag aatagtgtat gcggcgaccg agttgctctt
     9961 gcccggcgtc aatacgggat aataccgcgc cacatagcag aactttaaaa gtgctcatca
    10021 ttggaaaacg ttcttcgggg cgaaaactct caaggatctt accgctgttg agatccagtt
    10081 cgatgtaacc cactcgtgca cccaactgat cttcagcatc ttttactttc accagcgttt
    10141 ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa gggaataagg gcgacacgga
    10201 aatgttgaat actcatactc ttcctttttc aatattattg aagcatttat cagggttatt
    10261 gtctcatgag cggatacata tttgaatgta tttagaaaaa taaacaaata ggggttccgc
    10321 gcacatttcc ccgaaaagtg ccacctg
//
