pET28a/Cas9-Cys

LOCUS       pET28a/Cas9-Cys              9546 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Expresses N-terminal His tag fused human codon-optimized Cas9 
            nuclease having C-terminal Cysteine .
ACCESSION   .
VERSION     .
KEYWORDS    pET28a/Cas9-Cys
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9546)
  AUTHORS   Ramakrishna S, Kwaku Dad AB, Beloor J, Gopalappa R, Lee SK, Kim H
  TITLE     Gene disruption by cell-penetrating peptide-mediated delivery of 
            Cas9 protein and guide RNA.
  JOURNAL   Genome Res. 2014 Apr 2.
  PUBMED    24696462
REFERENCE   2  (bases 1 to 9546)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..9546
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     34..53
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        34..52
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    53..77
                     /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)."
     RBS             92..114
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             133..150
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             160..177
                     /codon_start=1
                     /product="thrombin recognition and cleavage site"
                     /label=thrombin site
                     /translation="LVPRGS"
     CDS             181..213
                     /codon_start=1
                     /product="leader peptide from bacteriophage T7 gene 10"
                     /label=T7 tag (gene 10 leader)
                     /note="promotes efficient translation in E. coli"
                     /translation="MASMTGGQQMG"
     CDS             256..4359
                     /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             4375..4395
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             4396..4422
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             4444..4461
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(4514..4532)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      4528..4575
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     rep_origin      4612..5067
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4699..4718)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4909..4930
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5159..5974)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     5882..5901
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     5966..5985
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      6096..6684
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6585..6604
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6838..6855
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    6870..7009
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(6935..6954)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7095..7117
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(7111..7302)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(8111..9193)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     9155..9174
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(9194..9271)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     9477..9496
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 gcgtagagga tcgagatctc gatcccgcga aattaatacg actcactata ggggaattgt
       61 gagcggataa caattcccct ctagaaataa ttttgtttaa ctttaagaag gagatatacc
      121 atgggcagca gccatcatca tcatcatcac agcagcggcc tggtgccgcg cggcagccat
      181 atggctagca tgactggtgg acagcaaatg ggtcgcggat ccgaattcga gctccgtcga
      241 caagcttgcg gccgcatgga caagaagtac agcatcggcc tggacatcgg taccaacagc
      301 gtgggctggg ccgtgatcac cgacgagtac aaggtgccca gcaagaagtt caaggtgctg
      361 ggcaacaccg accgccacag catcaagaag aacctgatcg gcgccctgct gttcgacagc
      421 ggcgagaccg ccgaggccac ccgcctgaag cgcaccgccc gccgccgcta cacccgccgc
      481 aagaaccgca tctgctacct gcaggagatc ttcagcaacg agatggccaa ggtggacgac
      541 agcttcttcc accgcctgga ggagagcttc ctggtggagg aggacaagaa gcacgagcgc
      601 caccccatct tcggcaacat cgtggacgag gtggcctacc acgagaagta ccccaccatc
      661 taccacctgc gcaagaagct ggtggacagc accgacaagg ccgacctgcg cctgatctac
      721 ctggccctgg cccacatgat caagttccgc ggccacttcc tgatcgaggg cgacctgaac
      781 cccgacaaca gcgacgtgga caagctgttc atccagctgg tgcagaccta caaccagctg
      841 ttcgaggaga accccatcaa cgccagcggc gtggacgcca aggccatcct gagcgcccgc
      901 ctgagcaaga gccgccgcct ggagaacctg atcgcccagc tgcccggcga gaagaagaac
      961 ggcctgttcg gcaacctgat cgccctgagc ctgggcctga cccccaactt caagagcaac
     1021 ttcgacctgg ccgaggacgc caagctgcag ctgagcaagg acacctacga cgacgacctg
     1081 gacaacctgc tggcccagat cggcgaccag tacgccgacc tgttcctggc cgccaagaac
     1141 ctgagcgacg ccatcctgct gagcgacatc ctgcgcgtga acaccgagat caccaaggcc
     1201 cccctgagcg ccagcatgat caagcgctac gacgagcacc accaggacct gaccctgctg
     1261 aaggccctgg tgcgccagca gctgcccgag aagtacaagg agatcttctt cgaccagagc
     1321 aagaacggct acgccggcta catcgacggc ggcgccagcc aggaggagtt ctacaagttc
     1381 atcaagccca tcctggagaa gatggacggc accgaggagc tgctggtgaa gctgaaccgc
     1441 gaggacctgc tgcgcaagca gcgcaccttc gacaacggca gcatccccca ccagatccac
     1501 ctgggcgagc tgcacgccat cctgcgccgc caggaggact tctacccctt cctgaaggac
     1561 aaccgcgaga agatcgagaa gatcctgacc ttccgcatcc cctactacgt gggccccctg
     1621 gcccgcggca acagccgctt cgcctggatg acccgcaaga gcgaggagac catcaccccc
     1681 tggaacttcg aggaggtggt ggacaagggc gccagcgccc agagcttcat cgagcgcatg
     1741 accaacttcg acaagaacct gcccaacgag aaggtgctgc ccaagcacag cctgctgtac
     1801 gagtacttca ccgtgtacaa cgagctgacc aaggtgaagt acgtgaccga gggcatgcgc
     1861 aagcccgcct tcctgagcgg cgagcagaag aaggccatcg tggacctgct gttcaagacc
     1921 aaccgcaagg tgaccgtgaa gcagctgaag gaggactact tcaagaagat cgagtgcttc
     1981 gacagcgtgg agatcagcgg cgtggaggac cgcttcaacg ccagcctggg cacctaccac
     2041 gacctgctga agatcatcaa ggacaaggac ttcctggaca acgaggagaa cgaggacatc
     2101 ctggaggaca tcgtgctgac cctgaccctg ttcgaggacc gcgagatgat cgaggagcgc
     2161 ctgaagacct acgcccacct gttcgacgac aaggtgatga agcagctgaa gcgccgccgc
     2221 tacaccggct ggggccgcct gagccgcaag cttatcaacg gcatccgcga caagcagagc
     2281 ggcaagacca tcctggactt cctgaagagc gacggcttcg ccaaccgcaa cttcatgcag
     2341 ctgatccacg acgacagcct gaccttcaag gaggacatcc agaaggccca ggtgagcggc
     2401 cagggcgaca gcctgcacga gcacatcgcc aacctggccg gcagccccgc catcaagaag
     2461 ggcatcctgc agaccgtgaa ggtggtggac gagctggtga aggtgatggg ccgccacaag
     2521 cccgagaaca tcgtgatcga gatggcccgc gagaaccaga ccacccagaa gggccagaag
     2581 aacagccgcg agcgcatgaa gcgcatcgag gagggcatca aggagctggg cagccagatc
     2641 ctgaaggagc accccgtgga gaacacccag ctgcagaacg agaagctgta cctgtactac
     2701 ctgcagaacg gccgcgacat gtacgtggac caggagctgg acatcaaccg cctgagcgac
     2761 tacgacgtgg accacatcgt gccccagagc ttcctgaagg acgacagcat cgacaacaag
     2821 gtgctgaccc gcagcgacaa gaaccgcggc aagagcgaca acgtgcccag cgaggaggtg
     2881 gtgaagaaga tgaagaacta ctggcgccag ctgctgaacg ccaagctgat cacccagcgc
     2941 aagttcgaca acctgaccaa ggccgagcgc ggcggcctga gcgagctgga caaggccggc
     3001 ttcatcaagc gccagctggt ggagacccgc cagatcacca agcacgtggc ccagatcctg
     3061 gacagccgca tgaacaccaa gtacgacgag aacgacaagc tgatccgcga ggtgaaggtg
     3121 atcaccctga agagcaagct ggtgagcgac ttccgcaagg acttccagtt ctacaaggtg
     3181 cgcgagatca acaactacca ccacgcccac gacgcctacc tgaacgccgt ggtgggcacc
     3241 gccctgatca agaagtaccc caagctggag agcgagttcg tgtacggcga ctacaaggtg
     3301 tacgacgtgc gcaagatgat cgccaagagc gagcaggaga tcggcaaggc caccgccaag
     3361 tacttcttct acagcaacat catgaacttc ttcaagaccg agatcaccct ggccaacggc
     3421 gagatccgca agcgccccct gatcgagacc aacggcgaga ccggcgagat cgtgtgggac
     3481 aagggccgcg acttcgccac cgtgcgcaag gtgctgagca tgccccaggt gaacatcgtg
     3541 aagaagaccg aggtgcagac cggcggcttc agcaaggaga gcatcctgcc caagcgcaac
     3601 agcgacaagc tgatcgcccg caagaaggac tgggacccca agaagtacgg cggcttcgac
     3661 agccccaccg tggcctacag cgtgctggtg gtggccaagg tggagaaggg caagagcaag
     3721 aagctgaaga gcgtgaagga gctgctgggc atcaccatca tggagcgcag cagcttcgag
     3781 aagaacccca tcgacttcct ggaggccaag ggctacaagg aggtgaagaa ggacctgatc
     3841 atcaagctgc ccaagtacag cctgttcgag ctggagaacg gccgcaagcg catgctggcc
     3901 agcgccggcg agctgcagaa gggcaacgag ctggccctgc ccagcaagta cgtgaacttc
     3961 ctgtacctgg ccagccacta cgagaagctg aagggcagcc ccgaggacaa cgagcagaag
     4021 cagctgttcg tggagcagca caagcactac ctggacgaga tcatcgagca gatcagcgag
     4081 ttcagcaagc gcgtgatcct ggccgacgcc aacctggaca aggtgctgag cgcctacaac
     4141 aagcaccgcg acaagcccat ccgcgagcag gccgagaaca tcatccacct gttcaccctg
     4201 accaacctgg gcgcccccgc cgccttcaag tacttcgaca ccaccatcga ccgcaagcgc
     4261 tacaccagca ccaaggaggt gctggacgcc accctgatcc accagagcat caccggtctg
     4321 tacgagaccc gcatcgacct gagccagctg ggcggcgacg gcggctccgg acctccaaag
     4381 aaaaagagaa aagtataccc ctacgacgtg cccgactacg cctgttaagc ggccgcactc
     4441 gagcaccacc accaccacca ctgagatccg gctgctaaca aagcccgaaa ggaagctgag
     4501 ttggctgctg ccaccgctga gcaataacta gcataacccc ttggggcctc taaacgggtc
     4561 ttgaggggtt ttttgctgaa aggaggaact atatccggat tggcgaatgg gacgcgccct
     4621 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     4681 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     4741 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     4801 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     4861 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     4921 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     4981 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     5041 ttaacaaaat attaacgctt acaatttagg tggcactttt cggggaaatg tgcgcggaac
     5101 ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga attaattctt
     5161 agaaaaactc atcgagcatc aaatgaaact gcaatttatt catatcagga ttatcaatac
     5221 catatttttg aaaaagccgt ttctgtaatg aaggagaaaa ctcaccgagg cagttccata
     5281 ggatggcaag atcctggtat cggtctgcga ttccgactcg tccaacatca atacaaccta
     5341 ttaatttccc ctcgtcaaaa ataaggttat caagtgagaa atcaccatga gtgacgactg
     5401 aatccggtga gaatggcaaa agtttatgca tttctttcca gacttgttca acaggccagc
     5461 cattacgctc gtcatcaaaa tcactcgcat caaccaaacc gttattcatt cgtgattgcg
     5521 cctgagcgag acgaaatacg cgatcgctgt taaaaggaca attacaaaca ggaatcgaat
     5581 gcaaccggcg caggaacact gccagcgcat caacaatatt ttcacctgaa tcaggatatt
     5641 cttctaatac ctggaatgct gttttcccgg ggatcgcagt ggtgagtaac catgcatcat
     5701 caggagtacg gataaaatgc ttgatggtcg gaagaggcat aaattccgtc agccagttta
     5761 gtctgaccat ctcatctgta acatcattgg caacgctacc tttgccatgt ttcagaaaca
     5821 actctggcgc atcgggcttc ccatacaatc gatagattgt cgcacctgat tgcccgacat
     5881 tatcgcgagc ccatttatac ccatataaat cagcatccat gttggaattt aatcgcggcc
     5941 tagagcaaga cgtttcccgt tgaatatggc tcataacacc ccttgtatta ctgtttatgt
     6001 aagcagacag ttttattgtt catgaccaaa atcccttaac gtgagttttc gttccactga
     6061 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     6121 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     6181 gagccaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     6241 gtccttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     6301 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     6361 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6421 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6481 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6541 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6601 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6661 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6721 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     6781 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     6841 gagtcagtga gcgaggaagc ggaagagcgc ctgatgcggt attttctcct tacgcatctg
     6901 tgcggtattt cacaccgcaa tggtgcactc tcagtacaat ctgctctgat gccgcatagt
     6961 taagccagta tacactccgc tatcgctacg tgactgggtc atggctgcgc cccgacaccc
     7021 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     7081 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     7141 cgcgaggcag ctgcggtaaa gctcatcagc gtggtcgtga agcgattcac agatgtctgc
     7201 ctgttcatcc gcgtccagct cgttgagttt ctccagaagc gttaatgtct ggcttctgat
     7261 aaagcgggcc atgttaaggg cggttttttc ctgtttggtc actgatgcct ccgtgtaagg
     7321 gggatttctg ttcatggggg taatgatacc gatgaaacga gagaggatgc tcacgatacg
     7381 ggttactgat gatgaacatg cccggttact ggaacgttgt gagggtaaac aactggcggt
     7441 atggatgcgg cgggaccaga gaaaaatcac tcagggtcaa tgccagcgct tcgttaatac
     7501 agatgtaggt gttccacagg gtagccagca gcatcctgcg atgcagatcc ggaacataat
     7561 ggtgcagggc gctgacttcc gcgtttccag actttacgaa acacggaaac cgaagaccat
     7621 tcatgttgtt gctcaggtcg cagacgtttt gcagcagcag tcgcttcacg ttcgctcgcg
     7681 tatcggtgat tcattctgct aaccagtaag gcaaccccgc cagcctagcc gggtcctcaa
     7741 cgacaggagc acgatcatgc gcacccgtgg ggccgccatg ccggcgataa tggcctgctt
     7801 ctcgccgaaa cgtttggtgg cgggaccagt gacgaaggct tgagcgaggg cgtgcaagat
     7861 tccgaatacc gcaagcgaca ggccgatcat cgtcgcgctc cagcgaaagc ggtcctcgcc
     7921 gaaaatgacc cagagcgctg ccggcacctg tcctacgagt tgcatgataa agaagacagt
     7981 cataagtgcg gcgacgatag tcatgccccg cgcccaccgg aaggagctga ctgggttgaa
     8041 ggctctcaag ggcatcggtc gagatcccgg tgcctaatga gtgagctaac ttacattaat
     8101 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     8161 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgccagggtg gtttttcttt
     8221 tcaccagtga gacgggcaac agctgattgc ccttcaccgc ctggccctga gagagttgca
     8281 gcaagcggtc cacgctggtt tgccccagca ggcgaaaatc ctgtttgatg gtggttaacg
     8341 gcgggatata acatgagctg tcttcggtat cgtcgtatcc cactaccgag atatccgcac
     8401 caacgcgcag cccggactcg gtaatggcgc gcattgcgcc cagcgccatc tgatcgttgg
     8461 caaccagcat cgcagtggga acgatgccct cattcagcat ttgcatggtt tgttgaaaac
     8521 cggacatggc actccagtcg ccttcccgtt ccgctatcgg ctgaatttga ttgcgagtga
     8581 gatatttatg ccagccagcc agacgcagac gcgccgagac agaacttaat gggcccgcta
     8641 acagcgcgat ttgctggtga cccaatgcga ccagatgctc cacgcccagt cgcgtaccgt
     8701 cttcatggga gaaaataata ctgttgatgg gtgtctggtc agagacatca agaaataacg
     8761 ccggaacatt agtgcaggca gcttccacag caatggcatc ctggtcatcc agcggatagt
     8821 taatgatcag cccactgacg cgttgcgcga gaagattgtg caccgccgct ttacaggctt
     8881 cgacgccgct tcgttctacc atcgacacca ccacgctggc acccagttga tcggcgcgag
     8941 atttaatcgc cgcgacaatt tgcgacggcg cgtgcagggc cagactggag gtggcaacgc
     9001 caatcagcaa cgactgtttg cccgccagtt gttgtgccac gcggttggga atgtaattca
     9061 gctccgccat cgccgcttcc actttttccc gcgttttcgc agaaacgtgg ctggcctggt
     9121 tcaccacgcg ggaaacggtc tgataagaga caccggcata ctctgcgaca tcgtataacg
     9181 ttactggttt cacattcacc accctgaatt gactctcttc cgggcgctat catgccatac
     9241 cgcgaaaggt tttgcgccat tcgatggtgt ccgggatctc gacgctctcc cttatgcgac
     9301 tcctgcatta ggaagcagcc cagtagtagg ttgaggccgt tgagcaccgc cgccgcaagg
     9361 aatggtgcat gcaaggagat ggcgcccaac agtcccccgg ccacggggcc tgccaccata
     9421 cccacgccga aacaagcgct catgagcccg aagtggcgag cccgatcttc cccatcggtg
     9481 atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg
     9541 cgtccg
//