pJSC011 - Bacterial expression plasmid for SpCas9 Cluster 1

LOCUS       pJSC011 - Bacterial expression plasmid for SpCas9 Cluster 1 (HypaCas9), HNH FRET variant               10073 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression plasmid for SpCas9 Cluster 1 (HypaCas9), HNH 
            FRET variant.
ACCESSION   .
VERSION     .
KEYWORDS   pJSC011 - Bacterial expression plasmid for SpCas9 Cluster 1 (HypaCas9), HNH FRET variant
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10073)
  AUTHORS   Chen JS, Dagdas YS, Kleinstiver BP, Welch MM, Sousa AA, Harrington 
            LB, Sternberg SH, Joung JK, Yildiz A, Doudna JA
  TITLE     Enhanced proofreading governs CRISPR-Cas9 targeting accuracy.
  JOURNAL   Nature. 2017 Sep 20. doi: 10.1038/nature24268.
  PUBMED    28931002
REFERENCE   2  (bases 1 to 10073)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..10073
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     1167..1186
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     1270..1289
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        1270..1288
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             1377..1399
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             1419..1448
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             1458..2558
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRIAATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     2532..2555
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             2613..2633
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2640..6746
                     /codon_start=1
                     /product="Cas9 endonuclease from the Streptococcus pyogenes
                     Type II CRISPR/Cas system, mutated to improve targeting 
                     specificity without compromising on-target activity (Chen 
                     et al., 2017)"
                     /label=HypaCas9
                     /note="carries the mutations N692A, M694A, Q695A, and 
                     H698A"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRISYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQCKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIESFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRAFA
                     ALIADDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKCDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     primer_bind     complement(6872..6890)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6886..6933
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(7299..7327)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(7354..7372)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7440..7544
                     /gene="bla"
                     /label=AmpR promoter
     CDS             7545..8405
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(7763..7782)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8576..9164
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9065..9084
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9318..9335
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    9350..9489
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(9415..9434)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     9575..9597
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(9591..9782)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ttacgaaaca cggaaaccga agaccattca tgttgttgct caggtcgcag acgttttgca
       61 gcagcagtcg cttcacgttc gctcgcgtat cggtgattca ttctgctaac cagtaaggca
      121 accccgccag cctagccggg tcctcaacga caggagcacg atcatgcgca cccgtggcca
      181 ggacccaacg ctgcccgaga tgcgccgcgt gcggctgctg gagatggcgg acgcgatgga
      241 tatgttctgc caagggttgg tttgcgcatt cacagttctc cgcaagaatt gattggctcc
      301 aattcttgga gtggtgaatc cgttagcgag gtgccgccgg cttccattca ggtcgaggtg
      361 gcccggctcc atgcaccgcg acgcaacgcg gggaggcaga caaggtatag ggcggcgcct
      421 acaatccatg ccaacccgtt ccatgtgctc gccgaggcgg cataaatcgc cgtgacgatc
      481 agcggtccaa tgatcgaagt taggctggta agagccgcga gcgatccttg aagctgtccc
      541 tgatggtcgt catctacctg cctggacagc atggcctgca acgcgggcat cccgatgccg
      601 ccggaagcga gaagaatcat aatggggaag gccatccagc ctcgcgtcgc gaacgcaagc
      661 aagacgtagc ccagcgcgtc ggccgccatg ccggcgataa tggcctgctt ctcgccgaaa
      721 cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat tccgaatacc
      781 gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc gaaaatgacc
      841 cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt cataagtgcg
      901 gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa ggctctcaag
      961 ggcatcggtc gacgctctcc cttatgcgac tcctgcatta ggaagcagcc cagtagtagg
     1021 ttgaggccgt tgagcaccgc cgccgcaagg aatggtgcat gcaaggagat ggcgcccaac
     1081 agtcccccgg ccacggggcc tgccaccata cccacgccga aacaagcgct catgagcccg
     1141 aagtggcgag cccgatcttc cccatcggtg atgtcggcga tataggcgcc agcaaccgca
     1201 cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt agaggatcga gatctcgatc
     1261 ccgcgaaatt aatacgactc actataggga gaccacaacg gtttccctct agtgccggct
     1321 ccggagagct ctttaattaa gcggccgccc tgcaggactc gagttctaga aataattttg
     1381 tttaacttta agaaggagat atacatatga aatcttctca ccatcaccat caccatcacc
     1441 atcaccatgg ttcttctatg aaaatcgaag aaggtaaact ggtaatctgg attaacggcg
     1501 ataaaggcta taacggtctc gctgaagtcg gtaagaaatt cgagaaagat accggaatta
     1561 aagtcaccgt tgagcatccg gataaactgg aagagaaatt cccacaggtt gcggcaactg
     1621 gcgatggccc tgacattatc ttctgggcac acgaccgctt tggtggctac gctcaatctg
     1681 gcctgttggc tgaaatcacc ccggacaaag cgttccagga caagctgtat ccgtttacct
     1741 gggatgccgt acgttacaac ggcaagctga ttgcttaccc gatcgctgtt gaagcgttat
     1801 cgctgattta taacaaagat ctgctgccga acccgccaaa aacctgggaa gagatcccgg
     1861 cgctggataa agaactgaaa gcgaaaggta agagcgcgct gatgttcaac ctgcaagaac
     1921 cgtacttcac ctggccgctg attgctgctg acgggggtta tgcgttcaag tatgaaaacg
     1981 gcaagtacga cattaaagac gtgggcgtgg ataacgctgg cgcaaaagcg ggtctgacct
     2041 tcctggttga cctgattaaa aacaaacaca tgaatgcaga caccgattac tccatcgcag
     2101 aagctgcctt taataaaggc gaaacagcga tgaccatcaa cggcccgtgg gcatggtcca
     2161 acatcgacac cagcaaagtg aattatggtg taacggtact gccgaccttc aagggtcaac
     2221 catccaaacc gttcgttggc gtgctgagcg caggtattaa cgccgccagt ccgaacaaag
     2281 agctggcaaa agagttcctc gaaaactatc tgctgactga tgaaggtctg gaagcggtta
     2341 ataaagacaa accgctgggt gccgtagcgc tgaagtctta cgaggaagag ttggcgaaag
     2401 atccacgtat tgccgccact atggaaaacg cccagaaagg tgaaatcatg ccgaacatcc
     2461 cgcagatgtc cgctttctgg tatgccgtgc gtactgcggt gatcaacgcc gccagcggtc
     2521 gtcagactgt cgatgaagcc ctgaaagacg cgcagactaa ttcgagctcg aacaacaaca
     2581 acaataacaa taacaacaac ctcgggatcg aggaaaacct gtacttccaa tccaatgcaa
     2641 tggataagaa atactcaata ggcttagata tcggcacaaa tagcgtcgga tgggcggtga
     2701 tcactgatga atataaggtt ccgtctaaaa agttcaaggt tctgggaaat acagaccgcc
     2761 acagtatcaa aaaaaatctt ataggggctc ttttatttga cagtggagag acagcggaag
     2821 cgactcgtct caaacggaca gctcgtagaa ggtatacacg tcggaagaat cgtatttctt
     2881 atctacagga gattttttca aatgagatgg cgaaagtaga tgatagtttc tttcatcgac
     2941 ttgaagagtc ttttttggtg gaagaagaca agaagcatga acgtcatcct atttttggaa
     3001 atatagtaga tgaagttgct tatcatgaga aatatccaac tatctatcat ctgcgaaaaa
     3061 aattggtaga ttctactgat aaagcggatt tgcgcttaat ctatttggcc ttagcgcata
     3121 tgattaagtt tcgtggtcat tttttgattg agggagattt aaatcctgat aatagtgatg
     3181 tggacaaact atttatccag ttggtacaaa cctacaatca attatttgaa gaaaacccta
     3241 ttaacgcaag tggagtagat gctaaagcga ttctttctgc acgattgagt aaatcaagac
     3301 gattagaaaa tctcattgct cagctccccg gtgagaagaa aaatggctta tttgggaatc
     3361 tcattgcttt gtcattgggt ttgaccccta attttaaatc aaattttgat ttggcagaag
     3421 atgctaaatt acagctttca aaagatactt acgatgatga tttagataat ttattggcgc
     3481 aaattggaga tcaatatgct gatttgtttt tggcagctaa gaatttatca gatgctattt
     3541 tactttcaga tatcctaaga gtaaatactg aaataactaa ggctccccta tcagcttcaa
     3601 tgattaaacg ctacgatgaa catcatcaag acttgactct tttaaaagct ttagttcgac
     3661 aacaacttcc agaaaagtat aaagaaatct tttttgatca atgtaaaaac ggatatgcag
     3721 gttatattga tgggggagct agccaagaag aattttataa atttatcaaa ccaattttag
     3781 aaaaaatgga tggtactgag gaattattgg tgaaactaaa tcgtgaagat ttgctgcgca
     3841 agcaacggac ctttgacaac ggctctattc cccatcaaat tcacttgggt gagctgcatg
     3901 ctattttgag aagacaagaa gacttttatc catttttaaa agacaatcgt gagaagattg
     3961 aaaaaatctt gacttttcga attccttatt atgttggtcc attggcgcgt ggcaatagtc
     4021 gttttgcatg gatgactcgg aagtctgaag aaacaattac cccatggaat tttgaagaag
     4081 ttgtcgataa aggtgcttca gctcaatcat ttattgaacg catgacaaac tttgataaaa
     4141 atcttccaaa tgaaaaagta ctaccaaaac atagtttgct ttatgagtat tttacggttt
     4201 ataacgaatt gacaaaggtc aaatatgtta ctgaaggaat gcgaaaacca gcatttcttt
     4261 caggtgaaca gaagaaagcc attgttgatt tactcttcaa aacaaatcga aaagtaaccg
     4321 ttaagcaatt aaaagaagat tatttcaaaa aaatagaatc ttttgatagt gttgaaattt
     4381 caggagttga agatagattt aatgcttcat taggtaccta ccatgatttg ctaaaaatta
     4441 ttaaagataa agattttttg gataatgaag aaaatgaaga tatcttagag gatattgttt
     4501 taacattgac cttatttgaa gatagggaga tgattgagga aagacttaaa acatatgctc
     4561 acctctttga tgataaggtg atgaaacagc ttaaacgtcg ccgttatact ggttggggac
     4621 gtttgtctcg aaaattgatt aatggtatta gggataagca atctggcaaa acaatattag
     4681 attttttgaa atcagatggt tttgccaatc gcgcttttgc tgctctgatc gctgatgata
     4741 gtttgacatt taaagaagac attcaaaaag cacaagtgtc tggacaaggc gatagtttac
     4801 atgaacatat tgcaaattta gctggtagcc ctgctattaa aaaaggtatt ttacagactg
     4861 taaaagttgt tgatgaattg gtcaaagtaa tggggcggca taagccagaa aatatcgtta
     4921 ttgaaatggc acgtgaaaat cagacaactc aaaagggcca gaaaaattcg cgagagcgta
     4981 tgaaacgaat cgaagaaggt atcaaagaat taggaagtca gattcttaaa gagcatcctg
     5041 ttgaaaatac tcaattgcaa aatgaaaagc tctatctcta ttatctccaa aatggaagag
     5101 acatgtatgt ggaccaagaa ttagatatta atcgtttaag tgattatgat gtcgatcaca
     5161 ttgttccaca aagtttcctt aaagacgatt caatagacaa taaggtctta acgcgttctg
     5221 ataaaaatcg tggtaaatgc gataacgttc caagtgaaga agtagtcaaa aagatgaaaa
     5281 actattggag acaacttcta aacgccaagt taatcactca acgtaagttt gataatttaa
     5341 cgaaagctga acgtggaggt ttgagtgaac ttgataaagc tggttttatc aaacgccaat
     5401 tggttgaaac tcgccaaatc actaagcatg tggcacaaat tttggatagt cgcatgaata
     5461 ctaaatacga tgaaaatgat aaacttattc gagaggttaa agtgattacc ttaaaatcta
     5521 aattagtttc tgacttccga aaagatttcc aattctataa agtacgtgag attaacaatt
     5581 accatcatgc ccatgatgcg tatctaaatg ccgtcgttgg aactgctttg attaagaaat
     5641 atccaaaact tgaatcggag tttgtctatg gtgattataa agtttatgat gttcgtaaaa
     5701 tgattgctaa gtctgagcaa gaaataggca aagcaaccgc aaaatatttc ttttactcta
     5761 atatcatgaa cttcttcaaa acagaaatta cacttgcaaa tggagagatt cgcaaacgcc
     5821 ctctaatcga aactaatggg gaaactggag aaattgtctg ggataaaggg cgagattttg
     5881 ccacagtgcg caaagtattg tccatgcccc aagtcaatat tgtcaagaaa acagaagtac
     5941 agacaggcgg attctccaag gagtcaattt taccaaaaag aaattcggac aagcttattg
     6001 ctcgtaaaaa agactgggat ccaaaaaaat atggtggttt tgatagtcca acggtagctt
     6061 attcagtcct agtggttgct aaggtggaaa aagggaaatc gaagaagtta aaatccgtta
     6121 aagagttact agggatcaca attatggaaa gaagttcctt tgaaaaaaat ccgattgact
     6181 ttttagaagc taaaggatat aaggaagtta aaaaagactt aatcattaaa ctacctaaat
     6241 atagtctttt tgagttagaa aacggtcgta aacggatgct ggctagtgcc ggagaattac
     6301 aaaaaggaaa tgagctggct ctgccaagca aatatgtgaa ttttttatat ttagctagtc
     6361 attatgaaaa gttgaagggt agtccagaag ataacgaaca aaaacaattg tttgtggagc
     6421 agcataagca ttatttagat gagattattg agcaaatcag tgaattttct aagcgtgtta
     6481 ttttagcaga tgccaattta gataaagttc ttagtgcata taacaaacat agagacaaac
     6541 caatacgtga acaagcagaa aatattattc atttatttac gttgacgaat cttggagctc
     6601 ccgctgcttt taaatatttt gatacaacaa ttgatcgtaa acgatatacg tctacaaaag
     6661 aagttttaga tgccactctt atccatcaat ccatcactgg tctttatgaa acacgcattg
     6721 atttgagtca gctaggaggt gactaataac attggaagtg gataacggat ccgcgatcgc
     6781 ggcgcgccac ctggtggccg gccggtacca cgcgtgcgct ctgatccggc tgctaacaaa
     6841 gcccgaaagg aagctgagtt ggctgctgcc accgctgagc aataactagc ataacccctt
     6901 ggggcctcta aacgggtctt gaggggtttt ttgctgaaag gaggaactat atccggatat
     6961 ccacaggacg ggtgtggtcg ccatgatcgc gtagtcgata gtggctccaa gtagcgaagc
     7021 gagcaggact gggcggcggc caaagcggtc ggacagtgct ccgagaacgg gtgcgcatag
     7081 aaattgcatc aacgcatata gcgctagcag cacgccatag tgactggcga tgctgtcgga
     7141 atggacgata tcccgcaaga ggcccggcag taccggcata accaagccta tgcctacagc
     7201 atccagggtg acggtgccga ggatgacgat gagcgcattg ttagatttca tacacggtgc
     7261 ctgactgcgt tagcaattta actgtgataa actaccgcat taaagcttat cgatgataag
     7321 ctgtcaaaca tgagaattct tgaagacgaa agggcctcgt gatacgccta tttttatagg
     7381 ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc
     7441 gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac
     7501 aataaccctg ataaatgctt caataacatt gaaaaaggaa gagtatgagt attcaacatt
     7561 tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag
     7621 aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg ggttacatcg
     7681 aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa cgttttccaa
     7741 tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtgtt gacgccgggc
     7801 aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag tactcaccag
     7861 tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt gctgccataa
     7921 ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga ccgaaggagc
     7981 taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt tgggaaccgg
     8041 agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgca gcaatggcaa
     8101 caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg caacaattaa
     8161 tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc cttccggctg
     8221 gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt atcattgcag
     8281 cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg gggagtcagg
     8341 caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg attaagcatt
     8401 ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa cttcattttt
     8461 aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa atcccttaac
     8521 gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag
     8581 atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg
     8641 tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact ggcttcagca
     8701 gagcgcagat accaaatact gtccttctag tgtagccgta gttaggccac cacttcaaga
     8761 actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg gctgctgcca
     8821 gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg gataaggcgc
     8881 agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga acgacctaca
     8941 ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc gaagggagaa
     9001 aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg agggagcttc
     9061 cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc
     9121 gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg
     9181 cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt cctgcgttat
     9241 cccctgattc tgtggataac cgtattaccg cctttgagtg agctgatacc gctcgccgca
     9301 gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt
     9361 attttctcct tacgcatctg tgcggtattt cacaccgcaa tggtgcactc tcagtacaat
     9421 ctgctctgat gccgcatagt taagccagta tacactccgc tatcgctacg tgactgggtc
     9481 atggctgcgc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc
     9541 ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg tcagaggttt
     9601 tcaccgtcat caccgaaacg cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga
     9661 agcgattcac agatgtctgc ctgttcatcc gcgtccagct cgttgagttt ctccagaagc
     9721 gttaatgtct ggcttctgat aaagcgggcc atgttaaggg cggttttttc ctgtttggtc
     9781 actgatgcct ccgtgtaagg gggatttctg ttcatggggg taatgatacc gatgaaacga
     9841 gagaggatgc tcacgatacg ggttactgat gatgaacatg cccggttact ggaacgttgt
     9901 gagggtaaac aactggcggt atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa
     9961 tgccagcgct tcgttaatac agatgtaggt gttccacagg gtagccagca gcatcctgcg
    10021 atgcagatcc ggaacataat ggtgcagggc gctgacttcc gcgtttccag act
//