pJSC282 - Bacterial expression plasmid for SpCas9 Cluster 1

LOCUS       pJSC282 - Bacterial expression plasmid for SpCas9 Cluster 1 (HypaCas9), REC3 FRET variant               10073 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression plasmid for SpCas9 Cluster 1 (HypaCas9), REC3 
            FRET variant.
ACCESSION   .
VERSION     .
KEYWORDS    pJSC282 - Bacterial expression plasmid for SpCas9 Cluster 1
            (H...
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.biofeng.com
FEATURES             Location/Qualifiers
     source          1..10073
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     170..189
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     273..292
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        273..291
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             380..402
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             422..451
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             461..1561
                     /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     1535..1558
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1616..1636
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1643..5749
                     /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
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIESFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRAFA
                     ALIADDCLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKCKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     primer_bind     complement(5875..5893)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5889..5936
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(6302..6330)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(6357..6375)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6443..6547
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6548..7408
                     /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(6766..6785)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7579..8167
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8068..8087
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     8321..8338
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    8353..8492
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(8418..8437)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8578..8600
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(8594..8785)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 ttaggaagca gcccagtagt aggttgaggc cgttgagcac cgccgccgca aggaatggtg
       61 catgcaagga gatggcgccc aacagtcccc cggccacggg gcctgccacc atacccacgc
      121 cgaaacaagc gctcatgagc ccgaagtggc gagcccgatc ttccccatcg gtgatgtcgg
      181 cgatataggc gccagcaacc gcacctgtgg cgccggtgat gccggccacg atgcgtccgg
      241 cgtagaggat cgagatctcg atcccgcgaa attaatacga ctcactatag ggagaccaca
      301 acggtttccc tctagtgccg gctccggaga gctctttaat taagcggccg ccctgcagga
      361 ctcgagttct agaaataatt ttgtttaact ttaagaagga gatatacata tgaaatcttc
      421 tcaccatcac catcaccatc accatcacca tggttcttct atgaaaatcg aagaaggtaa
      481 actggtaatc tggattaacg gcgataaagg ctataacggt ctcgctgaag tcggtaagaa
      541 attcgagaaa gataccggaa ttaaagtcac cgttgagcat ccggataaac tggaagagaa
      601 attcccacag gttgcggcaa ctggcgatgg ccctgacatt atcttctggg cacacgaccg
      661 ctttggtggc tacgctcaat ctggcctgtt ggctgaaatc accccggaca aagcgttcca
      721 ggacaagctg tatccgttta cctgggatgc cgtacgttac aacggcaagc tgattgctta
      781 cccgatcgct gttgaagcgt tatcgctgat ttataacaaa gatctgctgc cgaacccgcc
      841 aaaaacctgg gaagagatcc cggcgctgga taaagaactg aaagcgaaag gtaagagcgc
      901 gctgatgttc aacctgcaag aaccgtactt cacctggccg ctgattgctg ctgacggggg
      961 ttatgcgttc aagtatgaaa acggcaagta cgacattaaa gacgtgggcg tggataacgc
     1021 tggcgcaaaa gcgggtctga ccttcctggt tgacctgatt aaaaacaaac acatgaatgc
     1081 agacaccgat tactccatcg cagaagctgc ctttaataaa ggcgaaacag cgatgaccat
     1141 caacggcccg tgggcatggt ccaacatcga caccagcaaa gtgaattatg gtgtaacggt
     1201 actgccgacc ttcaagggtc aaccatccaa accgttcgtt ggcgtgctga gcgcaggtat
     1261 taacgccgcc agtccgaaca aagagctggc aaaagagttc ctcgaaaact atctgctgac
     1321 tgatgaaggt ctggaagcgg ttaataaaga caaaccgctg ggtgccgtag cgctgaagtc
     1381 ttacgaggaa gagttggcga aagatccacg tattgccgcc actatggaaa acgcccagaa
     1441 aggtgaaatc atgccgaaca tcccgcagat gtccgctttc tggtatgccg tgcgtactgc
     1501 ggtgatcaac gccgccagcg gtcgtcagac tgtcgatgaa gccctgaaag acgcgcagac
     1561 taattcgagc tcgaacaaca acaacaataa caataacaac aacctcggga tcgaggaaaa
     1621 cctgtacttc caatccaatg caatggataa gaaatactca ataggcttag atatcggcac
     1681 aaatagcgtc ggatgggcgg tgatcactga tgaatataag gttccgtcta aaaagttcaa
     1741 ggttctggga aatacagacc gccacagtat caaaaaaaat cttatagggg ctcttttatt
     1801 tgacagtgga gagacagcgg aagcgactcg tctcaaacgg acagctcgta gaaggtatac
     1861 acgtcggaag aatcgtattt cttatctaca ggagattttt tcaaatgaga tggcgaaagt
     1921 agatgatagt ttctttcatc gacttgaaga gtcttttttg gtggaagaag acaagaagca
     1981 tgaacgtcat cctatttttg gaaatatagt agatgaagtt gcttatcatg agaaatatcc
     2041 aactatctat catctgcgaa aaaaattggt agattctact gataaagcgg atttgcgctt
     2101 aatctatttg gccttagcgc atatgattaa gtttcgtggt cattttttga ttgagggaga
     2161 tttaaatcct gataatagtg atgtggacaa actatttatc cagttggtac aaacctacaa
     2221 tcaattattt gaagaaaacc ctattaacgc aagtggagta gatgctaaag cgattctttc
     2281 tgcacgattg agtaaatcaa gacgattaga aaatctcatt gctcagctcc ccggtgagaa
     2341 gaaaaatggc ttatttggga atctcattgc tttgtcattg ggtttgaccc ctaattttaa
     2401 atcaaatttt gatttggcag aagatgctaa attacagctt tcaaaagata cttacgatga
     2461 tgatttagat aatttattgg cgcaaattgg agatcaatat gctgatttgt ttttggcagc
     2521 taagaattta tcagatgcta ttttactttc agatatccta agagtaaata ctgaaataac
     2581 taaggctccc ctatcagctt caatgattaa acgctacgat gaacatcatc aagacttgac
     2641 tcttttaaaa gctttagttc gacaacaact tccagaaaag tataaagaaa tcttttttga
     2701 tcaatcaaaa aacggatatg caggttatat tgatggggga gctagccaag aagaatttta
     2761 taaatttatc aaaccaattt tagaaaaaat ggatggtact gaggaattat tggtgaaact
     2821 aaatcgtgaa gatttgctgc gcaagcaacg gacctttgac aacggctcta ttccccatca
     2881 aattcacttg ggtgagctgc atgctatttt gagaagacaa gaagactttt atccattttt
     2941 aaaagacaat cgtgagaaga ttgaaaaaat cttgactttt cgaattcctt attatgttgg
     3001 tccattggcg cgtggcaata gtcgttttgc atggatgact cggaagtctg aagaaacaat
     3061 taccccatgg aattttgaag aagttgtcga taaaggtgct tcagctcaat catttattga
     3121 acgcatgaca aactttgata aaaatcttcc aaatgaaaaa gtactaccaa aacatagttt
     3181 gctttatgag tattttacgg tttataacga attgacaaag gtcaaatatg ttactgaagg
     3241 aatgcgaaaa ccagcatttc tttcaggtga acagaagaaa gccattgttg atttactctt
     3301 caaaacaaat cgaaaagtaa ccgttaagca attaaaagaa gattatttca aaaaaataga
     3361 atcttttgat agtgttgaaa tttcaggagt tgaagataga tttaatgctt cattaggtac
     3421 ctaccatgat ttgctaaaaa ttattaaaga taaagatttt ttggataatg aagaaaatga
     3481 agatatctta gaggatattg ttttaacatt gaccttattt gaagataggg agatgattga
     3541 ggaaagactt aaaacatatg ctcacctctt tgatgataag gtgatgaaac agcttaaacg
     3601 tcgccgttat actggttggg gacgtttgtc tcgaaaattg attaatggta ttagggataa
     3661 gcaatctggc aaaacaatat tagatttttt gaaatcagat ggttttgcca atcgcgcttt
     3721 tgctgctctg atcgctgatg attgtttgac atttaaagaa gacattcaaa aagcacaagt
     3781 gtctggacaa ggcgatagtt tacatgaaca tattgcaaat ttagctggta gccctgctat
     3841 taaaaaaggt attttacaga ctgtaaaagt tgttgatgaa ttggtcaaag taatggggcg
     3901 gcataagcca gaaaatatcg ttattgaaat ggcacgtgaa aatcagacaa ctcaaaaggg
     3961 ccagaaaaat tcgcgagagc gtatgaaacg aatcgaagaa ggtatcaaag aattaggaag
     4021 tcagattctt aaagagcatc ctgttgaaaa tactcaattg caaaatgaaa agctctatct
     4081 ctattatctc caaaatggaa gagacatgta tgtggaccaa gaattagata ttaatcgttt
     4141 aagtgattat gatgtcgatc acattgttcc acaaagtttc cttaaagacg attcaataga
     4201 caataaggtc ttaacgcgtt ctgataaaaa tcgtggtaaa tcggataacg ttccaagtga
     4261 agaagtagtc aaaaagatga aaaactattg gagacaactt ctaaacgcca agttaatcac
     4321 tcaacgtaag tttgataatt taacgaaagc tgaacgtgga ggtttgagtg aacttgataa
     4381 agctggtttt atcaaacgcc aattggttga aactcgccaa atcactaagc atgtggcaca
     4441 aattttggat agtcgcatga atactaaata cgatgaaaat gataaactta ttcgagaggt
     4501 taaagtgatt accttaaaat gtaaattagt ttctgacttc cgaaaagatt tccaattcta
     4561 taaagtacgt gagattaaca attaccatca tgcccatgat gcgtatctaa atgccgtcgt
     4621 tggaactgct ttgattaaga aatatccaaa acttgaatcg gagtttgtct atggtgatta
     4681 taaagtttat gatgttcgta aaatgattgc taagtctgag caagaaatag gcaaagcaac
     4741 cgcaaaatat ttcttttact ctaatatcat gaacttcttc aaaacagaaa ttacacttgc
     4801 aaatggagag attcgcaaac gccctctaat cgaaactaat ggggaaactg gagaaattgt
     4861 ctgggataaa gggcgagatt ttgccacagt gcgcaaagta ttgtccatgc cccaagtcaa
     4921 tattgtcaag aaaacagaag tacagacagg cggattctcc aaggagtcaa ttttaccaaa
     4981 aagaaattcg gacaagctta ttgctcgtaa aaaagactgg gatccaaaaa aatatggtgg
     5041 ttttgatagt ccaacggtag cttattcagt cctagtggtt gctaaggtgg aaaaagggaa
     5101 atcgaagaag ttaaaatccg ttaaagagtt actagggatc acaattatgg aaagaagttc
     5161 ctttgaaaaa aatccgattg actttttaga agctaaagga tataaggaag ttaaaaaaga
     5221 cttaatcatt aaactaccta aatatagtct ttttgagtta gaaaacggtc gtaaacggat
     5281 gctggctagt gccggagaat tacaaaaagg aaatgagctg gctctgccaa gcaaatatgt
     5341 gaatttttta tatttagcta gtcattatga aaagttgaag ggtagtccag aagataacga
     5401 acaaaaacaa ttgtttgtgg agcagcataa gcattattta gatgagatta ttgagcaaat
     5461 cagtgaattt tctaagcgtg ttattttagc agatgccaat ttagataaag ttcttagtgc
     5521 atataacaaa catagagaca aaccaatacg tgaacaagca gaaaatatta ttcatttatt
     5581 tacgttgacg aatcttggag ctcccgctgc ttttaaatat tttgatacaa caattgatcg
     5641 taaacgatat acgtctacaa aagaagtttt agatgccact cttatccatc aatccatcac
     5701 tggtctttat gaaacacgca ttgatttgag tcagctagga ggtgactaat aacattggaa
     5761 gtggataacg gatccgcgat cgcggcgcgc cacctggtgg ccggccggta ccacgcgtgc
     5821 gcgctgatcc ggctgctaac aaagcccgaa aggaagctga gttggctgct gccaccgctg
     5881 agcaataact agcataaccc attggggcct ctaaacgggt cttgaggggt tttttgctga
     5941 aaggaggaac tatatccgga tatccacagg acgggtgtgg tcgccatgat cgcgtagtcg
     6001 atagtggctc caagtagcga agcgagcagg actgggcggc ggccaaagcg gtcggacagt
     6061 gctccgagaa cgggtgcgca tagaaattgc atcaacgcat atagcgctag cagcacgcca
     6121 tagtgactgg cgatgctgtc ggaatggacg atatcccgca agaggcccgg cagtaccggc
     6181 ataaccaagc ctatgcctac agcatccagg gtgacggtgc cgaggatgac gatgagcgca
     6241 ttgttagatt tcatacacgg tgcctgactg cgttagcaat ttaactgtga taaactaccg
     6301 cattaaagct tatcgatgat aagctgtcaa acatgagaat tcttgaagac gaaagggcct
     6361 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacgtcagg
     6421 tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct aaatacattc
     6481 aaatatgtat ccgctcatga gacaataacc ctgataaatg cttcaataac attgaaaaag
     6541 gaagagtatg agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg
     6601 ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt
     6661 gggtgcacga gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt
     6721 tcgccccgaa gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt
     6781 attatcccgt gttgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa
     6841 tgacttggtt gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag
     6901 agaattatgc agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac
     6961 aacgatcgga ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac
     7021 tcgccttgat cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac
     7081 cacgatgcct gcagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac
     7141 tctagcttcc cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact
     7201 tctgcgctcg gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg
     7261 tgggtctcgc ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt
     7321 tatctacacg acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat
     7381 aggtgcctca ctgattaagc attggtaact gtcagaccaa gtttactcat atatacttta
     7441 gattgattta aaacttcatt tttaatttaa aaggatctag gtgaagatcc tttttgataa
     7501 tctcatgacc aaaatccctt aacgtgagtt ttcgttccac tgagcgtcag accccgtaga
     7561 aaagatcaaa ggatcttctt gagatccttt ttttctgcgc gtaatctgct gcttgcaaac
     7621 aaaaaaacca ccgctaccag cggtggtttg tttgccggat caagagctac caactctttt
     7681 tccgaaggta actggcttca gcagagcgca gataccaaat actgtccttc tagtgtagcc
     7741 gtagttaggc caccacttca agaactctgt agcaccgcct acatacctcg ctctgctaat
     7801 cctgttacca gtggctgctg ccagtggcga taagtcgtgt cttaccgggt tggactcaag
     7861 acgatagtta ccggataagg cgcagcggtc gggctgaacg gggggttcgt gcacacagcc
     7921 cagcttggag cgaacgacct acaccgaact gagataccta cagcgtgagc tatgagaaag
     7981 cgccacgctt cccgaaggga gaaaggcgga caggtatccg gtaagcggca gggtcggaac
     8041 aggagagcgc acgagggagc ttccaggggg aaacgcctgg tatctttata gtcctgtcgg
     8101 gtttcgccac ctctgacttg agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct
     8161 atggaaaaac gccagcaacg cggccttttt acggttcctg gccttttgct ggccttttgc
     8221 tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta ccgcctttga
     8281 gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag tgagcgagga
     8341 agcggaagag cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     8401 caatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gtatacactc
     8461 cgctatcgct acgtgactgg gtcatggctg cgccccgaca cccgccaaca cccgctgacg
     8521 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     8581 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgagg cagctgcggt
     8641 aaagctcatc agcgtggtcg tgaagcgatt cacagatgtc tgcctgttca tccgcgtcca
     8701 gctcgttgag tttctccaga agcgttaatg tctggcttct gataaagcgg gccatgttaa
     8761 gggcggtttt ttcctgtttg gtcactgatg cctccgtgta agggggattt ctgttcatgg
     8821 gggtaatgat accgatgaaa cgagagagga tgctcacgat acgggttact gatgatgaac
     8881 atgcccggtt actggaacgt tgtgagggta aacaactggc ggtatggatg cggcgggacc
     8941 agagaaaaat cactcagggt caatgccagc gcttcgttaa tacagatgta ggtgttccac
     9001 agggtagcca gcagcatcct gcgatgcaga tccggaacat aatggtgcag ggcgctgact
     9061 tccgcgtttc cagactttac gaaacacgga aaccgaagac cattcatgtt gttgctcagg
     9121 tcgcagacgt tttgcagcag cagtcgcttc acgttcgctc gcgtatcggt gattcattct
     9181 gctaaccagt aaggcaaccc cgccagccta gccgggtcct caacgacagg agcacgatca
     9241 tgcgcacccg tggccaggac ccaacgctgc ccgagatgcg ccgcgtgcgg ctgctggaga
     9301 tggcggacgc gatggatatg ttctgccaag ggttggtttg cgcattcaca gttctccgca
     9361 agaattgatt ggctccaatt cttggagtgg tgaatccgtt agcgaggtgc cgccggcttc
     9421 cattcaggtc gaggtggccc ggctccatgc accgcgacgc aacgcgggga ggcagacaag
     9481 gtatagggcg gcgcctacaa tccatgccaa cccgttccat gtgctcgccg aggcggcata
     9541 aatcgccgtg acgatcagcg gtccaatgat cgaagttagg ctggtaagag ccgcgagcga
     9601 tccttgaagc tgtccctgat ggtcgtcatc tacctgcctg gacagcatgg cctgcaacgc
     9661 gggcatcccg atgccgccgg aagcgagaag aatcataatg gggaaggcca tccagcctcg
     9721 cgtcgcgaac gccagcaaga cgtagcccag cgcgtcggcc gccatgccgg cgataatggc
     9781 ctgcttctcg ccgaaacgtt tggtggcggg accagtgacg aaggcttgag cgagggcgtg
     9841 caagattccg aataccgcaa gcgacaggcc gatcatcgtc gcgctccagc gaaagcggtc
     9901 ctcgccgaaa atgacccaga gcgctgccgg cacctgtcct acgagttgca tgataaagaa
     9961 gacagtcata agtgcggcga cgatagtcat gccccgcgcc caccggaagg agctgactgg
    10021 gttgaaggct ctcaagggca tcggtcgacg ctctccctta tgcgactcct gca
//