pJSC115 - Bacterial expression plasmid for SpCas9-HF1, HNH F

LOCUS       pJSC115 - Bacterial expression plasmid for SpCas9-HF1, HNH FRET variant               10073 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Bacterial expression plasmid for SpCas9-HF1, HNH FRET variant.
ACCESSION   .
VERSION     .
KEYWORDS    pJSC115 - Bacterial expression plasmid for SpCas9-HF1, HNH FRE...
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     1016..1039
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             1097..1117
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             1124..5230
                     /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
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRISYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQCKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTAFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIESFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGALSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     ALIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKCDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRAITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     primer_bind     complement(5356..5374)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5370..5417
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(5783..5811)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(5838..5856)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5924..6028
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6029..6889
                     /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(6247..6266)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7060..7648
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7549..7568
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7802..7819
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    7834..7973
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(7899..7918)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     8059..8081
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(8075..8266)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     9724..9743
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     9827..9846
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        9827..9845
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             9934..9956
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             9976..10005
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             10015..1042
                     /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"
ORIGIN
        1 tctcgctgaa gtcggtaaga aattcgagaa agataccgga attaaagtca ccgttgagca
       61 tccggataaa ctggaagaga aattcccaca ggttgcggca actggcgatg gccctgacat
      121 tatcttctgg gcacacgacc gctttggtgg ctacgctcaa tctggcctgt tggctgaaat
      181 caccccggac aaagcgttcc aggacaagct gtatccgttt acctgggatg ccgtacgtta
      241 caacggcaag ctgattgctt acccgatcgc tgttgaagcg ttatcgctga tttataacaa
      301 agatctgctg ccgaacccgc caaaaacctg ggaagagatc ccggcgctgg ataaagaact
      361 gaaagcgaaa ggtaagagcg cgctgatgtt caacctgcaa gaaccgtact tcacctggcc
      421 gctgattgct gctgacgggg gttatgcgtt caagtatgaa aacggcaagt acgacattaa
      481 agacgtgggc gtggataacg ctggcgcaaa agcgggtctg accttcctgg ttgacctgat
      541 taaaaacaaa cacatgaatg cagacaccga ttactccatc gcagaagctg cctttaataa
      601 aggcgaaaca gcgatgacca tcaacggccc gtgggcatgg tccaacatcg acaccagcaa
      661 agtgaattat ggtgtaacgg tactgccgac cttcaagggt caaccatcca aaccgttcgt
      721 tggcgtgctg agcgcaggta ttaacgccgc cagtccgaac aaagagctgg caaaagagtt
      781 cctcgaaaac tatctgctga ctgatgaagg tctggaagcg gttaataaag acaaaccgct
      841 gggtgccgta gcgctgaagt cttacgagga agagttggcg aaagatccac gtattgccgc
      901 cactatggaa aacgcccaga aaggtgaaat catgccgaac atcccgcaga tgtccgcttt
      961 ctggtatgcc gtgcgtactg cggtgatcaa cgccgccagc ggtcgtcaga ctgtcgatga
     1021 agccctgaaa gacgcgcaga ctaattcgag ctcgaacaac aacaacaata acaataacaa
     1081 caacctcggg atcgaggaaa acctgtactt ccaatccaat gcaatggata agaaatactc
     1141 aataggctta gatatcggca caaatagcgt cggatgggcg gtgatcactg atgaatataa
     1201 ggttccgtct aaaaagttca aggttctggg aaatacagac cgccacagta tcaaaaaaaa
     1261 tcttataggg gctcttttat ttgacagtgg agagacagcg gaagcgactc gtctcaaacg
     1321 gacagctcgt agaaggtata cacgtcggaa gaatcgtatt tcttatctac aggagatttt
     1381 ttcaaatgag atggcgaaag tagatgatag tttctttcat cgacttgaag agtctttttt
     1441 ggtggaagaa gacaagaagc atgaacgtca tcctattttt ggaaatatag tagatgaagt
     1501 tgcttatcat gagaaatatc caactatcta tcatctgcga aaaaaattgg tagattctac
     1561 tgataaagcg gatttgcgct taatctattt ggccttagcg catatgatta agtttcgtgg
     1621 tcattttttg attgagggag atttaaatcc tgataatagt gatgtggaca aactatttat
     1681 ccagttggta caaacctaca atcaattatt tgaagaaaac cctattaacg caagtggagt
     1741 agatgctaaa gcgattcttt ctgcacgatt gagtaaatca agacgattag aaaatctcat
     1801 tgctcagctc cccggtgaga agaaaaatgg cttatttggg aatctcattg ctttgtcatt
     1861 gggtttgacc cctaatttta aatcaaattt tgatttggca gaagatgcta aattacagct
     1921 ttcaaaagat acttacgatg atgatttaga taatttattg gcgcaaattg gagatcaata
     1981 tgctgatttg tttttggcag ctaagaattt atcagatgct attttacttt cagatatcct
     2041 aagagtaaat actgaaataa ctaaggctcc cctatcagct tcaatgatta aacgctacga
     2101 tgaacatcat caagacttga ctcttttaaa agctttagtt cgacaacaac ttccagaaaa
     2161 gtataaagaa atcttttttg atcaatgtaa aaacggatat gcaggttata ttgatggggg
     2221 agctagccaa gaagaatttt ataaatttat caaaccaatt ttagaaaaaa tggatggtac
     2281 tgaggaatta ttggtgaaac taaatcgtga agatttgctg cgcaagcaac ggacctttga
     2341 caacggctct attccccatc aaattcactt gggtgagctg catgctattt tgagaagaca
     2401 agaagacttt tatccatttt taaaagacaa tcgtgagaag attgaaaaaa tcttgacttt
     2461 tcgaattcct tattatgttg gtccattggc gcgtggcaat agtcgttttg catggatgac
     2521 tcggaagtct gaagaaacaa ttaccccatg gaattttgaa gaagttgtcg ataaaggtgc
     2581 ttcagctcaa tcatttattg aacgcatgac agcgtttgat aaaaatcttc caaatgaaaa
     2641 agtactacca aaacatagtt tgctttatga gtattttacg gtttataacg aattgacaaa
     2701 ggtcaaatat gttactgaag gaatgcgaaa accagcattt ctttcaggtg aacagaagaa
     2761 agccattgtt gatttactct tcaaaacaaa tcgaaaagta accgttaagc aattaaaaga
     2821 agattatttc aaaaaaatag aatcttttga tagtgttgaa atttcaggag ttgaagatag
     2881 atttaatgct tcattaggta cctaccatga tttgctaaaa attattaaag ataaagattt
     2941 tttggataat gaagaaaatg aagatatctt agaggatatt gttttaacat tgaccttatt
     3001 tgaagatagg gagatgattg aggaaagact taaaacatat gctcacctct ttgatgataa
     3061 ggtgatgaaa cagcttaaac gtcgccgtta tactggttgg ggagcgttgt ctcgaaaatt
     3121 gattaatggt attagggata agcaatctgg caaaacaata ttagattttt tgaaatcaga
     3181 tggttttgcc aatcgcaatt ttatggccct gatccatgat gatagtttga catttaaaga
     3241 agacattcaa aaagcacaag tgtctggaca aggcgatagt ttacatgaac atattgcaaa
     3301 tttagctggt agccctgcta ttaaaaaagg tattttacag actgtaaaag ttgttgatga
     3361 attggtcaaa gtaatggggc ggcataagcc agaaaatatc gttattgaaa tggcacgtga
     3421 aaatcagaca actcaaaagg gccagaaaaa ttcgcgagag cgtatgaaac gaatcgaaga
     3481 aggtatcaaa gaattaggaa gtcagattct taaagagcat cctgttgaaa atactcaatt
     3541 gcaaaatgaa aagctctatc tctattatct ccaaaatgga agagacatgt atgtggacca
     3601 agaattagat attaatcgtt taagtgatta tgatgtcgat cacattgttc cacaaagttt
     3661 ccttaaagac gattcaatag acaataaggt cttaacgcgt tctgataaaa atcgtggtaa
     3721 atgcgataac gttccaagtg aagaagtagt caaaaagatg aaaaactatt ggagacaact
     3781 tctaaacgcc aagttaatca ctcaacgtaa gtttgataat ttaacgaaag ctgaacgtgg
     3841 aggtttgagt gaacttgata aagctggttt tatcaaacgc caattggttg aaactcgcgc
     3901 catcactaag catgtggcac aaattttgga tagtcgcatg aatactaaat acgatgaaaa
     3961 tgataaactt attcgagagg ttaaagtgat taccttaaaa tctaaattag tttctgactt
     4021 ccgaaaagat ttccaattct ataaagtacg tgagattaac aattaccatc atgcccatga
     4081 tgcgtatcta aatgccgtcg ttggaactgc tttgattaag aaatatccaa aacttgaatc
     4141 ggagtttgtc tatggtgatt ataaagttta tgatgttcgt aaaatgattg ctaagtctga
     4201 gcaagaaata ggcaaagcaa ccgcaaaata tttcttttac tctaatatca tgaacttctt
     4261 caaaacagaa attacacttg caaatggaga gattcgcaaa cgccctctaa tcgaaactaa
     4321 tggggaaact ggagaaattg tctgggataa agggcgagat tttgccacag tgcgcaaagt
     4381 attgtccatg ccccaagtca atattgtcaa gaaaacagaa gtacagacag gcggattctc
     4441 caaggagtca attttaccaa aaagaaattc ggacaagctt attgctcgta aaaaagactg
     4501 ggatccaaaa aaatatggtg gttttgatag tccaacggta gcttattcag tcctagtggt
     4561 tgctaaggtg gaaaaaggga aatcgaagaa gttaaaatcc gttaaagagt tactagggat
     4621 cacaattatg gaaagaagtt cctttgaaaa aaatccgatt gactttttag aagctaaagg
     4681 atataaggaa gttaaaaaag acttaatcat taaactacct aaatatagtc tttttgagtt
     4741 agaaaacggt cgtaaacgga tgctggctag tgccggagaa ttacaaaaag gaaatgagct
     4801 ggctctgcca agcaaatatg tgaatttttt atatttagct agtcattatg aaaagttgaa
     4861 gggtagtcca gaagataacg aacaaaaaca attgtttgtg gagcagcata agcattattt
     4921 agatgagatt attgagcaaa tcagtgaatt ttctaagcgt gttattttag cagatgccaa
     4981 tttagataaa gttcttagtg catataacaa acatagagac aaaccaatac gtgaacaagc
     5041 agaaaatatt attcatttat ttacgttgac gaatcttgga gctcccgctg cttttaaata
     5101 ttttgataca acaattgatc gtaaacgata tacgtctaca aaagaagttt tagatgccac
     5161 tcttatccat caatccatca ctggtcttta tgaaacacgc attgatttga gtcagctagg
     5221 aggtgactaa taacattgga agtggataac ggatccgcga tcgcggcgcg ccacctggtg
     5281 gccggccggt accacgcgtg cgctctgatc cggctgctaa caaagcccga aaggaagctg
     5341 agttggctgc tgccaccgct gagcaataac tagcataacc ccttggggcc tctaaacggg
     5401 tcttgagggg ttttttgctg aaaggaggaa ctatatccgg atatccacag gacgggtgtg
     5461 gtcgccatga tcgcgtagtc gatagtggct ccaagtagcg aagcgagcag gactgggcgg
     5521 cggccaaagc ggtcggacag tgctccgaga acgggtgcgc atagaaattg catcaacgca
     5581 tatagcgcta gcagcacgcc atagtgactg gcgatgctgt cggaatggac gatatcccgc
     5641 aagaggcccg gcagtaccgg cataaccaag cctatgccta cagcatccag ggtgacggtg
     5701 ccgaggatga cgatgagcgc attgttagat ttcatacacg gtgcctgact gcgttagcaa
     5761 tttaactgtg ataaactacc gcattaaagc ttatcgatga taagctgtca aacatgagaa
     5821 ttcttgaaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     5881 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     5941 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     6001 gcttcaataa cattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     6061 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     6121 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     6181 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     6241 agttctgcta tgtggcgcgg tattatcccg tgttgacgcc gggcaagagc aactcggtcg
     6301 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     6361 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     6421 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     6481 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     6541 accaaacgac gagcgtgaca ccacgatgcc tgcagcaatg gcaacaacgt tgcgcaaact
     6601 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6661 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6721 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     6781 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     6841 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     6901 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     6961 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     7021 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     7081 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     7141 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     7201 tactgtcctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     7261 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     7321 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     7381 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     7441 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     7501 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     7561 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     7621 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     7681 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     7741 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     7801 cagcgagtca gtgagcgagg aagcggaaga gcgcctgatg cggtattttc tccttacgca
     7861 tctgtgcggt atttcacacc gcaatggtgc actctcagta caatctgctc tgatgccgca
     7921 tagttaagcc agtatacact ccgctatcgc tacgtgactg ggtcatggct gcgccccgac
     7981 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     8041 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     8101 aacgcgcgag gcagctgcgg taaagctcat cagcgtggtc gtgaagcgat tcacagatgt
     8161 ctgcctgttc atccgcgtcc agctcgttga gtttctccag aagcgttaat gtctggcttc
     8221 tgataaagcg ggccatgtta agggcggttt tttcctgttt ggtcactgat gcctccgtgt
     8281 aagggggatt tctgttcatg ggggtaatga taccgatgaa acgagagagg atgctcacga
     8341 tacgggttac tgatgatgaa catgcccggt tactggaacg ttgtgagggt aaacaactgg
     8401 cggtatggat gcggcgggac cagagaaaaa tcactcaggg tcaatgccag cgcttcgtta
     8461 atacagatgt aggtgttcca cagggtagcc agcagcatcc tgcgatgcag atccggaaca
     8521 taatggtgca gggcgctgac ttccgcgttt ccagacttta cgaaacacgg aaaccgaaga
     8581 ccattcatgt tgttgctcag gtcgcagacg ttttgcagca gcagtcgctt cacgttcgct
     8641 cgcgtatcgg tgattcattc tgctaaccag taaggcaacc ccgccagcct agccgggtcc
     8701 tcaacgacag gagcacgatc atgcgcaccc gtggccagga cccaacgctg cccgagatgc
     8761 gccgcgtgcg gctgctggag atggcggacg cgatggatat gttctgccaa gggttggttt
     8821 gcgcattcac agttctccgc aagaattgat tggctccaat tcttggagtg gtgaatccgt
     8881 tagcgaggtg ccgccggctt ccattcaggt cgaggtggcc cggctccatg caccgcgacg
     8941 caacgcgggg aggcagacaa ggtatagggc ggcgcctaca atccatgcca acccgttcca
     9001 tgtgctcgcc gaggcggcat aaatcgccgt gacgatcagc ggtccaatga tcgaagttag
     9061 gctggtaaga gccgcgagcg atccttgaag ctgtccctga tggtcgtcat ctacctgcct
     9121 ggacagcatg gcctgcaacg cgggcatccc gatgccgccg gaagcgagaa gaatcataat
     9181 ggggaaggcc atccagcctc gcgtcgcgaa cgccagcaag acgtagccca gcgcgtcggc
     9241 cgccatgccg gcgataatgg cctgcttctc gccgaaacgt ttggtggcgg gaccagtgac
     9301 gaaggcttga gcgagggcgt gcaagattcc gaataccgca agcgacaggc cgatcatcgt
     9361 cgcgctccag cgaaagcggt cctcgccgaa aatgacccag agcgctgccg gcacctgtcc
     9421 tacgagttgc atgataaaga agacagtcat aagtgcggcg acgatagtca tgccccgcgc
     9481 ccaccggaag gagctgactg ggttgaaggc tctcaagggc atcggtcgac gctctccctt
     9541 atgcgactcc tgcattagga agcagcccag tagtaggttg aggccgttga gcaccgccgc
     9601 cgcaaggaat ggtgcatgca aggagatggc gcccaacagt cccccggcca cggggcctgc
     9661 caccataccc acgccgaaac aagcgctcat gagcccgaag tggcgagccc gatcttcccc
     9721 atcggtgatg tcggcgatat aggcgccagc aaccgcacct gtggcgccgg tgatgccggc
     9781 cacgatgcgt ccggcgtaga ggatcgagat ctcgatcccg cgaaattaat acgactcact
     9841 atagggagac cacaacggtt tccctctagt gccggctccg gagagctctt taattaagcg
     9901 gccgccctgc aggactcgag ttctagaaat aattttgttt aactttaaga aggagatata
     9961 catatgaaat cttctcacca tcaccatcac catcaccatc accatggttc ttctatgaaa
    10021 atcgaagaag gtaaactggt aatctggatt aacggcgata aaggctataa cgg
//