LOCUS       sequence-396878-        9728 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Thermosensitive pSC101-based plasmid expressing a sgRNA under the 
            control of the J23119 promoter and Cas9 under the control of the 
            DAPG-inducible PhlF promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9728)
  AUTHORS   Rostain W, Grebert T, Vyhovskyi D, Pizarro PT, Tshinsele-Van 
            Bellingen G, Cui L, Bikard D
  TITLE     Cas9 off-target binding to the promoter of bacterial genes leads to 
            silencing and toxicity.
  JOURNAL   Nucleic Acids Res. 2023 Mar 17:gkad170. doi: 10.1093/nar/gkad170.
  PUBMED    36929199
REFERENCE   2  (bases 1 to 9728)
  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: "Nucleic 
            Acids Res. 2023 Mar 17:gkad170. doi: 10.1093/nar/gkad170."
COMMENT     Sequence Label: pTG131
FEATURES             Location/Qualifiers
     source          1..9728
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(1..951)
                     /codon_start=1
                     /gene="rep101 encoding an A56V mutation"
                     /product="temperature-sensitive version of the RepA protein
                     needed for replication with the pSC101 origin (Armstrong et
                     al., 1984)"
                     /label=Rep101(Ts)
                     /translation="MSELVVFKANELAISRYDLTEHETKLILCCVALLNPTIENPTRKE
                     RTVSFTYNQYVQMMNISRENAYGVLAKATRELMTRTVEIRNPLVKGFEIFQWTNYAKFS
                     SEKLELVFSEEILPYLFQLKKFIKYNLEHVKSFENKYSMRIYEWLLKELTQKKTHKANI
                     EISLDEFKFMLMLENNYHEFKRLNQWVLKPISKDLNTYSNMKLVVDKRGRPTDTLIFQV
                     ELDRQMDLVTELENNQIKMNGDKIPTTITSDSYLHNGLRKTLHDALTAKIQLTSFEAKF
                     LSDMQSKYDLNGSFSWLTQKQRTTLENILAKYGRI"
     rep_origin      999..1221
                     /label=pSC101 ori
                     /note="low-copy replication origin that requires the Rep101
                     protein"
     CDS             complement(join(1875..2666,2667..2735))
                     /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     2498..2517
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2736..2840)
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3285..3319
                     /label=BBa_J23110
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.33"
     RBS             3329..3340
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             3972..3983
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     RBS             4091..4102
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             4109..8215
                     /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             4109..8212
                     /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"
     terminator      8284..8528
                     /label=lambda tL3 terminator
                     /note="transcription terminator tL3 from phage lambda"
     misc_RNA        complement(8572..8647)
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     terminator      complement(8702..8729)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        complement(9547..9581)
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
ORIGIN
        1 tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg gttcgttgtt tttgcgtgag
       61 ccatgagaac gaaccattga gatcatactt actttgcatg tcactcaaaa attttgcctc
      121 aaaactggtg agctgaattt ttgcagttaa agcatcgtgt agtgtttttc ttagtccgtt
      181 atgtaggtag gaatctgatg taatggttgt tggtattttg tcaccattca tttttatctg
      241 gttgttctca agttcggtta cgagatccat ttgtctatct agttcaactt ggaaaatcaa
      301 cgtatcagtc gggcggcctc gcttatcaac caccaatttc atattgctgt aagtgtttaa
      361 atctttactt attggtttca aaacccattg gttaagcctt ttaaactcat ggtagttatt
      421 ttcaagcatt aacatgaact taaattcatc aaggctaatc tctatatttg ccttgtgagt
      481 tttcttttgt gttagttctt ttaataacca ctcataaatc ctcatagagt atttgttttc
      541 aaaagactta acatgttcca gattatattt tatgaatttt tttaactgga aaagataagg
      601 caatatctct tcactaaaaa ctaattctaa tttttcgctt gagaacttgg catagtttgt
      661 ccactggaaa atctcaaagc ctttaaccaa aggattcctg atttccacag ttctcgtcat
      721 cagctctctg gttgctttag ctaatacacc ataagcattt tccctactga tgttcatcat
      781 ctgaacgtat tggttataag tgaacgatac cgtccgttct ttccttgtag ggttttcaat
      841 cgtggggttg agtagtgcca cacagcataa aattagcttg gtttcatgct ccgttaagtc
      901 atagcgacta atcgctagtt catttgcttt gaaaacaact aattcagaca tacatctcaa
      961 ttggtctagg tgattttaat cactatacca attgagatgg gctagtcaat gataattact
     1021 agtccttttc ctttgagttg tgggtatctg taaattctgc tagacctttg ctggaaaact
     1081 tgtaaattct gctagaccct ctgtaaattc cgctagacct ttgtgtgttt tttttgttta
     1141 tattcaagtg gttataattt atagaataaa gaaagaataa aaaaagataa aaagaataga
     1201 tcccagccct gtgtataact cactacttta gtcagttccg cagtattaca aaaggatgtc
     1261 gcaaacgctg tttgctcctc tacaaaacag accttaaaac cctaaaggct taagtagcac
     1321 cctcgcaagc tcggttgcgg ccgcaatcgg gcaaatcgct gaatattcct tttgtctccg
     1381 accatcaggc acctgagtcg ctgtcttttt cgtgacattc agttcgctgc gctcacggct
     1441 ctggcagtga atgggggtaa atggcactac aggcgccttt tatggattca tgcaaggaaa
     1501 ctacccataa tacaagaaaa gcccgtcacg ggcttctcag ggcgttttat ggcgggtctg
     1561 ctatgtggtg ctatctgact ttttgctgtt cagcagttcc tgccctctga ttttccagtc
     1621 tgaccacttc ggattatccc gtgacaggtc attcagactg gctaatgcac ccagtaaggc
     1681 agcggtatca tcaacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt
     1741 ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt
     1801 taaatcaatc taaagtatat atgagtaaac ttggtctgac agttacgttt ccaaccaatt
     1861 aaccaattct gattttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     1921 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     1981 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     2041 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     2101 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     2161 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     2221 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     2281 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     2341 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     2401 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     2461 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     2521 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     2581 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     2641 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     2701 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     2761 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     2821 aaacaaatag gggttccgcg aacacccctt gtattactgt ttatgtaagc agacagtttt
     2881 attgttcatg atgatatatt tttatcttgt gcaatgtaac atcagagatt ttgagacaca
     2941 acgtggcttt ccctgcagga tttcggaggc ctgcgttatc ccctgattct gtggataacc
     3001 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgccc caaaaagcct
     3061 cgctttcagc acctgtcgtt tcctttcttt tcagagggta ttttaaataa aaacattaag
     3121 ttatgacgaa gaagaacgga aacgccttaa accggaaaat tttcataaat agcgaaaacc
     3181 cgcgaggtcg ccgccccgta acctgtcgga tcaccggaaa ggacccgtaa agtgataatg
     3241 attatcatct acatatcaca acgtgcgtgg agggactagt ggattttacg gctagctcag
     3301 tcctaggtac aatgctagcg aattcattaa agaggagaaa ggtacccatg gcacgtaccc
     3361 cgagccgtag cagcattggt agcctgcgta gtccgcatac ccataaagca attctgacca
     3421 gcaccattga aatcctgaaa gaatgtggtt atagcggtct gagcattgaa agcgttgcac
     3481 gtcgtgccgg tgcaagcaaa ccgaccattt atcgttggtg gaccaataaa gcagcactga
     3541 ttgccgaagt gtatgaaaat gaaagcgaac aggtgcgtaa atttccggat ctgggtagct
     3601 ttaaagccga tctggatttt ctgctgcgta atctgtggaa agtttggcgt gaaaccattt
     3661 gtggtgaagc atttcgttgt gttattgcag aagcacagct ggaccctgca accctgaccc
     3721 agctgaaaga tcagtttatg gaacgtcgtc gtgagatgcc gaaaaaactg gttgaaaatg
     3781 ccattagcaa tggtgaactg ccgaaagata ccaatcgtga actgctgctg gatatgattt
     3841 ttggtttttg ttggtatcgc ctgctgaccg aacagctgac cgttgaacag gatattgaag
     3901 aatttacctt cctgctgatt aatggtgttt gtccgggtac acagcgttaa ctagggccca
     3961 tacccccaat tattgaaggc cgctaacgcg gccttttttt gtttctggtc tgcccgacgt
     4021 acggtgaatc tgattcgtta ccaattgaca tgatacgaaa cgtaccgtat cgttaaggtt
     4081 actagagatt aaagaggaga aatactagat ggataagaaa tactcaatag gcttagatat
     4141 cggcacaaat agcgtcggat gggcggtgat cactgatgaa tataaggttc cgtctaaaaa
     4201 gttcaaggtt ctgggaaata cagaccgcca cagtatcaaa aaaaatctta taggggctct
     4261 tttatttgac agtggagaga cagcggaagc gactcgtctc aaacggacag ctcgtagaag
     4321 gtatacacgt cggaagaatc gtatttgtta tctacaggag attttttcaa atgagatggc
     4381 gaaagtagat gatagtttct ttcatcgact tgaagagtct tttttggtgg aagaagacaa
     4441 gaagcatgaa cgtcatccta tttttggaaa tatagtagat gaagttgctt atcatgagaa
     4501 atatccaact atctatcatc tgcgaaaaaa attggtagat tctactgata aagcggattt
     4561 gcgcttaatc tatttggcct tagcgcatat gattaagttt cgtggtcatt ttttgattga
     4621 gggagattta aatcctgata atagtgatgt ggacaaacta tttatccagt tggtacaaac
     4681 ctacaatcaa ttatttgaag aaaaccctat taacgcaagt ggagtagatg ctaaagcgat
     4741 tctttctgca cgattgagta aatcaagacg attagaaaat ctcattgctc agctccccgg
     4801 tgagaagaaa aatggcttat ttgggaatct cattgctttg tcattgggtt tgacccctaa
     4861 ttttaaatca aattttgatt tggcagaaga tgctaaatta cagctttcaa aagatactta
     4921 cgatgatgat ttagataatt tattggcgca aattggagat caatatgctg atttgttttt
     4981 ggcagctaag aatttatcag atgctatttt actttcagat atcctaagag taaatactga
     5041 aataactaag gctcccctat cagcttcaat gattaaacgc tacgatgaac atcatcaaga
     5101 cttgactctt ttaaaagctt tagttcgaca acaacttcca gaaaagtata aagaaatctt
     5161 ttttgatcaa tcaaaaaacg gatatgcagg ttatattgat gggggagcta gccaagaaga
     5221 attttataaa tttatcaaac caattttaga aaaaatggat ggtactgagg aattattggt
     5281 gaaactaaat cgtgaagatt tgctgcgcaa gcaacggacc tttgacaacg gctctattcc
     5341 ccatcaaatt cacttgggtg agctgcatgc tattttgaga agacaagaag acttttatcc
     5401 atttttaaaa gacaatcgtg agaagattga aaaaatcttg acttttcgaa ttccttatta
     5461 tgttggtcca ttggcgcgtg gcaatagtcg ttttgcatgg atgactcgga agtctgaaga
     5521 aacaattacc ccatggaatt ttgaagaagt tgtcgataaa ggtgcttcag ctcaatcatt
     5581 tattgaacgc atgacaaact ttgataaaaa tcttccaaat gaaaaagtac taccaaaaca
     5641 tagtttgctt tatgagtatt ttacggttta taacgaattg acaaaggtca aatatgttac
     5701 tgaaggaatg cgaaaaccag catttctttc aggtgaacag aagaaagcca ttgttgattt
     5761 actcttcaaa acaaatcgaa aagtaaccgt taagcaatta aaagaagatt atttcaaaaa
     5821 aatagaatgt tttgatagtg ttgaaatttc aggagttgaa gatagattta atgcttcatt
     5881 aggtacctac catgatttgc taaaaattat taaagataaa gattttttgg ataatgaaga
     5941 aaatgaagat atcttagagg atattgtttt aacattgacc ttatttgaag atagggagat
     6001 gattgaggaa agacttaaaa catatgctca cctctttgat gataaggtga tgaaacagct
     6061 taaacgtcgc cgttatactg gttggggacg tttgtctcga aaattgatta atggtattag
     6121 ggataagcaa tctggcaaaa caatattaga ttttttgaaa tcagatggtt ttgccaatcg
     6181 caattttatg cagctgatcc atgatgatag tttgacattt aaagaagaca ttcaaaaagc
     6241 acaagtgtct ggacaaggcg atagtttaca tgaacatatt gcaaatttag ctggtagccc
     6301 tgctattaaa aaaggtattt tacagactgt aaaagttgtt gatgaattgg tcaaagtaat
     6361 ggggcggcat aagccagaaa atatcgttat tgaaatggca cgtgaaaatc agacaactca
     6421 aaagggccag aaaaattcgc gagagcgtat gaaacgaatc gaagaaggta tcaaagaatt
     6481 aggaagtcag attcttaaag agcatcctgt tgaaaatact caattgcaaa atgaaaagct
     6541 ctatctctat tatctccaaa atggaagaga catgtatgtg gaccaagaat tagatattaa
     6601 tcgtttaagt gattatgatg tcgatcacat tgttccacaa agtttcctta aagacgattc
     6661 aatagacaat aaggtcttaa cgcgttctga taaaaatcgt ggtaaatcgg ataacgttcc
     6721 aagtgaagaa gtagtcaaaa agatgaaaaa ctattggaga caacttctaa acgccaagtt
     6781 aatcactcaa cgtaagtttg ataatttaac gaaagctgaa cgtggaggtt tgagtgaact
     6841 tgataaagct ggttttatca aacgccaatt ggttgaaact cgccaaatca ctaagcatgt
     6901 ggcacaaatt ttggatagtc gcatgaatac taaatacgat gaaaatgata aacttattcg
     6961 agaggttaaa gtgattacct taaaatctaa attagtttct gacttccgaa aagatttcca
     7021 attctataaa gtacgtgaga ttaacaatta ccatcatgcc catgatgcgt atctaaatgc
     7081 cgtcgttgga actgctttga ttaagaaata tccaaaactt gaatcggagt ttgtctatgg
     7141 tgattataaa gtttatgatg ttcgtaaaat gattgctaag tctgagcaag aaataggcaa
     7201 agcaaccgca aaatatttct tttactctaa tatcatgaac ttcttcaaaa cagaaattac
     7261 acttgcaaat ggagagattc gcaaacgccc tctaatcgaa actaatgggg aaactggaga
     7321 aattgtctgg gataaagggc gagattttgc cacagtgcgc aaagtattgt ccatgcccca
     7381 agtcaatatt gtcaagaaaa cagaagtaca gacaggcgga ttctccaagg agtcaatttt
     7441 accaaaaaga aattcggaca agcttattgc tcgtaaaaaa gactgggatc caaaaaaata
     7501 tggtggtttt gatagtccaa cggtagctta ttcagtccta gtggttgcta aggtggaaaa
     7561 agggaaatcg aagaagttaa aatccgttaa agagttacta gggatcacaa ttatggaaag
     7621 aagttccttt gaaaaaaatc cgattgactt tttagaagct aaaggatata aggaagttaa
     7681 aaaagactta atcattaaac tacctaaata tagtcttttt gagttagaaa acggtcgtaa
     7741 acggatgctg gctagtgccg gagaattaca aaaaggaaat gagctggctc tgccaagcaa
     7801 atatgtgaat tttttatatt tagctagtca ttatgaaaag ttgaagggta gtccagaaga
     7861 taacgaacaa aaacaattgt ttgtggagca gcataagcat tatttagatg agattattga
     7921 gcaaatcagt gaattttcta agcgtgttat tttagcagat gccaatttag ataaagttct
     7981 tagtgcatat aacaaacata gagacaaacc aatacgtgaa caagcagaaa atattattca
     8041 tttatttacg ttgacgaatc ttggagctcc cgctgctttt aaatattttg atacaacaat
     8101 tgatcgtaaa cgatatacgt ctacaaaaga agttttagat gccactctta tccatcaatc
     8161 catcactggt ctttatgaaa cacgcattga tttgagtcag ctaggaggtg actaactcga
     8221 gtaaggatct ccaggcattg caggcatgcc tcgagatgca tggcgcctaa cctaaactga
     8281 tgacgcatcc tcacgataat atccgggtag gcgcaatcac tttcgtctac tccgttacaa
     8341 agcgaggctg ggtatttccc ggcctttctg ttatccgaaa tccactgaaa gcacagcggc
     8401 tggctgagga gataaataat aaacgagggg ctgtatgcac aaagcatctt ctgttgagtt
     8461 aagaacgagt atcgagatgg cacatagcct tgctcaaatt ggaatcaggt ttgtgccaat
     8521 accagtagaa acagacgaag aatccatggg tatggacaga tctcaaaaaa agcaccgact
     8581 cggtgccact ttttcaagtt gataacggac tagccttatt ttaacttgct atttctagct
     8641 ctaaaaccga gacctaactc aacaggtaag gcgcgaggtt ttttcaattg ttcgaaacgc
     8701 aaaaaggcca tccgtcagga tggccttctg cttaatttga tgcctgtcag tttaggttag
     8761 gcgccatgca tctcgaggca tgcctgcatt atttgtacag ctcatccatg ccaccggtag
     8821 aatgacgacc ctccgcgcgc tcatattgct ctacgatcgt ataatcttca ttatgagagg
     8881 tgatgtccag tttaatattc acattgtacg cgccaggcag ctgcacaggt ttcttggctt
     8941 tgtacgtggt tttcacttca gcgtcataat ggccgccatc tttcagtttc aggcgctgtt
     9001 taatttcgcc tttcagcgca ccatcttccg gatacatgcg ttcgctagac gcctcccaac
     9061 ccatcgtctt cttctgcatc accgggccat cagatggaaa attagtacca cgcagtttaa
     9121 ctttatagat gaactcgcca tcctgcaggg aggagtcctg agtgacggtc acgacaccac
     9181 catcttcaaa attcattacg cgttcccact tgaaaccttc cggaaaagac agcttcagat
     9241 agtccgggat atccgctggg tgtttaacat aagctttaga accgtacata aattgcgggc
     9301 tcaggatgtc ccacgcaaaa ggcagcgggc cgcctttagt cactttcagt ttggcggtct
     9361 gggtgccttc atacggacgg ccctcgcctt cgccttcgat ctcgaactcg tgaccgttaa
     9421 cagaaccctc catgtgaact ttgaagcgca tgaactcttt aatgatagcc atgttatcct
     9481 cctcgccctt ggaaaccatc tagatttctc ctttgtatca aaccatcacc cacgggactg
     9541 gtctcaacta gtattatacc taggactgag ctagctgtca accattcgat ggtgtcaacg
     9601 taaatgcatg ccgctcgcct tccatgggta tggacagttt tccctttgat atgtaacggt
     9661 gaacagttgt tctacttttg tttgttagtc ttgatgcttc actgatagat acaagagcca
     9721 taagaacc
//
