pH-PABE-7-esgRNA

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相关技术服务:质粒构建    基因合成    质粒大提

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载体基本信息

质粒名称: pH-PABE-7-esgRNA 
出品公司: 爱迪基因
目录编号: 115620
存储实验室: Caixia Gao

载体骨架基本信息
载体名称: pCAMBIA1300
载体出品公司: --
载体抗性: 卡那霉素
空载体大小: --
完整质粒大小: --
载体修饰: --
载体类型: 植物表达,CRISPR
筛选标记: --
高拷贝/低拷贝: 高拷贝
生长条件: 37 ℃

插入基因信息
插入基因名称: wtTadA-TadA7.10-nCas9-3*NLS
物种来源: --
插入大小: --
基因突变: --
GenBank编号 --
启动子 --
标签/融合蛋白: --
克隆方法: 限制性内切酶
5' 克隆位点: --
3' 克隆位点: --
5' 测序引物 --
3' 测序引物 --

文献引用方法
材料和方法部分: pH-PABE-7-esgRNA was a gift from Caixia Gao (Addgene plasmid # 115620 ; http://n2t.net/addgene:115620 ; RRID:Addgene_115620)
参考文献部分: Expanded base editing in rice and wheat using a Cas9-adenosine deaminase fusion. Li C, Zong Y, Wang Y, Jin S, Zhang D, Song Q, Zhang R, Gao C. Genome Biol. 2018 May 29;19(1):59. doi: 10.1186/s13059-018-1443-z. 10.1186/s13059-018-1443-z PubMed 29807545

载体图谱质粒图谱和多克隆位点信息

pH-PABE-7-esgRNA质粒图谱

载体简介

载体序列

LOCUS       pH-PABE-7-esgRNA               18869 bp ds-DNA     circular SYN 11-JAN-2019
DEFINITION  Targeted A to G in rice.
ACCESSION   .
VERSION     .
KEYWORDS    pH-PABE-7-esgRNA
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 18869)
  AUTHORS   Li C, Zong Y, Wang Y, Jin S, Zhang D, Song Q, Zhang R, Gao C
  TITLE     Expanded base editing in rice and wheat using a Cas9-adenosine 
            deaminase fusion.
  JOURNAL   Genome Biol. 2018 May 29;19(1):59. doi: 10.1186/s13059-018-1443-z.
  PUBMED    29807545
REFERENCE   2  (bases 1 to 18869)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 11, 2019 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..18869
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     misc_feature    261..285
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     primer_bind     473..495
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     487..504
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     488..504
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        571..950
                     /label=OsU3 promoter
                     /note="Oryza sativa (rice) snRNA U3 promoter"
     terminator      1058..1077
                     /label=SUP4 terminator
                     /note="transcription terminator for the S. cerevisiae SUP4 
                     tRNA gene"
     promoter        1090..3080
                     /label=Ubi promoter
                     /note="maize polyubiquitin gene promoter"
     CDS             4288..8388
                     /codon_start=1
                     /product="nickase mutant of the Cas9 endonuclease from the 
                     Streptococcus pyogenes Type II CRISPR/Cas system"
                     /label=Cas9(D10A)
                     /note="generates RNA-guided single strand nicks in DNA due 
                     to the D10A mutation in the RuvC catalytic domain"
                     /translation="DKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKKN
                     LIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEESF
                     LVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIKF
                     RGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRLE
                     NLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQI
                     GDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVRQ
                     QLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLLR
                     KQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARGN
                     SRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEYF
                     TVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFDS
                     VEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEERL
                     KTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFMQ
                     LIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGRH
                     KPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLYL
                     YYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVPS
                     EEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKHV
                     AQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYLN
                     AVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKTE
                     ITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSKE
                     SILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGIT
                     IMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNEL
                     ALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILADA
                     NLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVLD
                     ATLIHQSITGLYETRIDLSQLGGD"
     RBS             8301..8309
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     CDS             8401..8421
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8434..8454
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             8467..8487
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     primer_bind     complement(9146..9162)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(9146..9162)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(9159..9181)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    9170..9186
                     /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)."
     promoter        complement(9194..9224)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9239..9260
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        9451..10127
                     /label=CaMV 35S promoter (enhanced)
                     /note="cauliflower mosaic virus 35S promoter with a 
                     duplicated enhancer region"
     primer_bind     10064..10084
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             10194..11219
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRSQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     K"
     polyA_signal    11260..11434
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     misc_feature    11512..11536
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     CDS             11961..12755
                     /codon_start=1
                     /gene="aphA-3"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin"
                     /translation="MAKMRISPELKKLIEKYRCVKDTEGMSPAKVYKLVGENENLYLKM
                     TDSRYKGTTYDVEREKDMMLWLEGKLPVPKVLHFERHDGWSNLLMSEADGVLCSEEYED
                     EQSPEKIIELYAECIRLFHSIDISDCPYTNSLDSRLAELDYLLNNDLADVDCENWEEDT
                     PFKDPRELYDFLKTEKPEEELVFSHGDLGDSNIFVKDGKVSGFIDLGRSGRADKWYDIA
                     FCVRSIREDIGEEQYVELFFDLLGIKPDWEKIKYYILLDELF"
     rep_origin      14180..14768
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     14669..14688
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     14922..14939
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    14954..15094
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(15020..15039)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     15180..15202
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     rep_origin      15438..15632
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     CDS             complement(15698..16771)
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     CDS             complement(17200..17829)
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
ORIGIN
        1 gcgacaagaa aacgccagga aaagggcagg gcggcagcct gtcgcgtaac ttaggacttg
       61 tgcgacatgt cgttttcaga agacggctgc actgaacgtc agaagccgac tgcactatag
      121 cagcggaggg gttggatcaa agtactttga tcccgagggg aaccctgtgg ttggcatgca
      181 catacaaatg gacgaacgga taaacctttt cacgcccttt taaatatccg attattctaa
      241 taaacgctct tttctcttag gtttacccgc caatatatcc tgtcaaacac tgatagttta
      301 aactgaaggc gggaaacgac aatctgatcc aagctcaagc tgctctagca ttcgccattc
      361 aggctgcgca actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccagctg
      421 gcgaaagggg gatgtgctgc aaggcgatta agttgggtaa cgccagggtt ttcccagtca
      481 cgacgttgta aaacgacggc cagtgccaag cttagtaatt catccaggtc accaagttct
      541 aggattttca gaactgcaac ttattttatc aaggaatctt taaacatacg aacagatcac
      601 ttaaagttct tctgaagcaa cttaaagtta tcaggcatgc atggatcttg gaggaatcag
      661 atgtgcagtc agggaccata gcacaagaca ggcgtcttct actggtgcta ccagcaaatg
      721 ctggaagccg ggaacactgg gtacgttgga aaccacgtga tgtgaagaag taagataaac
      781 tgtaggagaa aagcatttcg tagtgggcca tgaagccttt caggacatgt attgcagtat
      841 gggccggccc attacgcaat tggacgacaa caaagactag tattagtacc acctcggcta
      901 tccacataga tcaaagctga tttaaaagag ttgtgcagat gatccgtggc gtgagaccct
      961 gcagggtctc tgtttaagag ctatgctgga aacagcatag caagtttaaa taaggctagt
     1021 ccgttatcaa cttgaaaaag tggcaccgag tcggtgcttt ttttgttttt tatgtctaag
     1081 cttgcatgcc tgcagtgcag cgtgacccgg tcgtgcccct ctctagagat aatgagcatt
     1141 gcatgtctaa gttataaaaa attaccacat attttttttg tcacacttgt ttgaagtgca
     1201 gtttatctat ctttatacat atatttaaac tttactctac gaataatata atctatagta
     1261 ctacaataat atcagtgttt tagagaatca tataaatgaa cagttagaca tggtctaaag
     1321 gacaattgag tattttgaca acaggactct acagttttat ctttttagtg tgcatgtgtt
     1381 ctcctttttt tttgcaaata gcttcaccta tataatactt catccatttt attagtacat
     1441 ccatttaggg tttagggtta atggttttta tagactaatt tttttagtac atctatttta
     1501 ttctatttta gcctctaaat taagaaaact aaaactctat tttagttttt ttatttaata
     1561 atttagatat aaaatagaat aaaataaagt gactaaaaat taaacaaata ccctttaaga
     1621 aattaaaaaa actaaggaaa catttttctt gtttcgagta gataatgcca gcctgttaaa
     1681 cgccgtcgac gagtctaacg gacaccaacc agcgaaccag cagcgtcgcg tcgggccaag
     1741 cgaagcagac ggcacggcat ctctgtcgct gcctctggac ccctctcgag agttccgctc
     1801 caccgttgga cttgctccgc tgtcggcatc cagaaattgc gtggcggagc ggcagacgtg
     1861 agccggcacg gcaggcggcc tcctcctcct ctcacggcac cggcagctac gggggattcc
     1921 tttcccaccg ctccttcgct ttcccttcct cgcccgccgt aataaataga caccccctcc
     1981 acaccctctt tccccaacct cgtgttgttc ggagcgcaca cacacacaac cagatctccc
     2041 ccaaatccac ccgtcggcac ctccgcttca aggtacgccg ctcgtcctcc ccccccccct
     2101 ctctaccttc tctagatcgg cgttccggtc catggttagg gcccggtagt tctacttctg
     2161 ttcatgtttg tgttagatcc gtgtttgtgt tagatccgtg ctgctagcgt tcgtacacgg
     2221 atgcgacctg tacgtcagac acgttctgat tgctaacttg ccagtgtttc tctttgggga
     2281 atcctgggat ggctctagcc gttccgcaga cgggatcgat ttcatgattt tttttgtttc
     2341 gttgcatagg gtttggtttg cccttttcct ttatttcaat atatgccgtg cacttgtttg
     2401 tcgggtcatc ttttcatgct tttttttgtc ttggttgtga tgatgtggtc tggttgggcg
     2461 gtcgttctag atcggagtag aattctgttt caaactacct ggtggattta ttaattttgg
     2521 atctgtatgt gtgtgccata catattcata gttacgaatt gaagatgatg gatggaaata
     2581 tcgatctagg ataggtatac atgttgatgc gggttttact gatgcatata cagagatgct
     2641 ttttgttcgc ttggttgtga tgatgtggtg tggttgggcg gtcgttcatt cgttctagat
     2701 cggagtagaa tactgtttca aactacctgg tgtatttatt aattttggaa ctgtatgtgt
     2761 gtgtcataca tcttcatagt tacgagttta agatggatgg aaatatcgat ctaggatagg
     2821 tatacatgtt gatgtgggtt ttactgatgc atatacatga tggcatatgc agcatctatt
     2881 catatgctct aaccttgagt acctatctat tataataaac aagtatgttt tataattatt
     2941 ttgatcttga tatacttgga tgatggcata tgcagcagct atatgtggat ttttttagcc
     3001 ctgccttcat acgctattta tttgcttggt actgtttctt ttgtcgatgc tcaccctgtt
     3061 gtttggtgtt acttctgcag ccctaggcct atgtccgagg tggagttctc tcacgagtat
     3121 tggatgaggc acgctcttac acttgctaag agagcttggg acgaaagaga agtgccagtt
     3181 ggcgccgttc ttgtgcataa taatagggtg atcggcgagg gttggaatag accaattgga
     3241 aggcatgatc caacagctca cgcagagatt atggctctca gacaaggcgg cctcgttatg
     3301 cagaactaca ggctcattga cgctacactc tacgtgacac tcgaaccttg cgttatgtgc
     3361 gccggagcta tgattcattc taggattggc agggtcgtgt ttggagctag ggacgctaaa
     3421 acaggagccg ccggatctct tatggacgtg ttgcatcatc caggcatgaa ccatagggtg
     3481 gagattacag agggcattct tgcagacgag tgcgctgctc ttctttccga tttcttcagg
     3541 atgagaaggc aggagattaa ggcccagaag aaggctcagt cttctacaga tagcggagga
     3601 tcttccggag gatctagcgg ctccgagaca ccaggaacat ccgaaagcgc tacaccagaa
     3661 tctagcggag gctcttccgg aggatcttct gaagtggagt tctcccacga gtattggatg
     3721 aggcacgctc ttacacttgc taaaagggct agggacgaaa gggaagttcc agttggagct
     3781 gttctcgtgc tcaataacag ggtgattggc gagggttgga atagagccat tggactccat
     3841 gatccaacag ctcacgcaga gattatggct cttagacaag gcggcctcgt tatgcagaat
     3901 tacagactca tcgacgccac actctacgtt accttcgaac cttgcgttat gtgcgccgga
     3961 gctatgatcc attctaggat tggcagggtc gtgttcggcg ttagaaacgc taagacagga
     4021 gctgcaggct ctcttatgga cgttcttcat tacccaggca tgaatcatag agtggagatc
     4081 acagaaggca ttcttgcaga cgagtgcgca gctctccttt gctatttctt caggatgccg
     4141 aggcaagttt tcaacgctca gaagaaggcc cagtcttcta cagattccgg cggatcttct
     4201 ggaggatcta gcggctccga gacaccagga acatccgaat ccgctacacc agagtcttct
     4261 ggaggatcta gcggaggatc tcttaaggac aagaagtact cgatcggcct cgccattggg
     4321 actaactctg ttggctgggc cgtgatcacc gacgagtaca aggtgccctc aaagaagttc
     4381 aaggtcctgg gcaacaccga tcggcattcc atcaagaaga atctcattgg cgctctcctg
     4441 ttcgacagcg gcgagacggc tgaggctacg cggctcaagc gcaccgcccg caggcggtac
     4501 acgcgcagga agaatcgcat ctgctacctg caggagattt tctccaacga gatggcgaag
     4561 gttgacgatt ctttcttcca caggctggag gagtcattcc tcgtggagga ggataagaag
     4621 cacgagcggc atccaatctt cggcaacatt gtcgacgagg ttgcctacca cgagaagtac
     4681 cctacgatct accatctgcg gaagaagctc gtggactcca cagataaggc ggacctccgc
     4741 ctgatctacc tcgctctggc ccacatgatt aagttcaggg gccatttcct gatcgagggg
     4801 gatctcaacc cggacaatag cgatgttgac aagctgttca tccagctcgt gcagacgtac
     4861 aaccagctct tcgaggagaa ccccattaat gcgtcaggcg tcgacgcgaa ggctatcctg
     4921 tccgctaggc tctcgaagtc tcggcgcctc gagaacctga tcgcccagct gccgggcgag
     4981 aagaagaacg gcctgttcgg gaatctcatt gcgctcagcc tggggctcac gcccaacttc
     5041 aagtcgaatt tcgatctcgc tgaggacgcc aagctgcagc tctccaagga cacatacgac
     5101 gatgacctgg ataacctcct ggcccagatc ggcgatcagt acgcggacct gttcctcgct
     5161 gccaagaatc tgtcggacgc catcctcctg tctgatattc tcagggtgaa caccgagatt
     5221 acgaaggctc cgctctcagc ctccatgatc aagcgctacg acgagcacca tcaggatctg
     5281 accctcctga aggcgctggt caggcagcag ctccccgaga agtacaagga gatcttcttc
     5341 gatcagtcga agaacggcta cgctgggtac attgacggcg gggcctctca ggaggagttc
     5401 tacaagttca tcaagccgat tctggagaag atggacggca cggaggagct gctggtgaag
     5461 ctcaatcgcg aggacctcct gaggaagcag cggacattcg ataacggcag catcccacac
     5521 cagattcatc tcggggagct gcacgctatc ctgaggaggc aggaggactt ctaccctttc
     5581 ctcaaggata accgcgagaa gatcgagaag attctgactt tcaggatccc gtactacgtc
     5641 ggcccactcg ctaggggcaa ctcccgcttc gcttggatga cccgcaagtc agaggagacg
     5701 atcacgccgt ggaacttcga ggaggtggtc gacaagggcg ctagcgctca gtcgttcatc
     5761 gagaggatga cgaatttcga caagaacctg ccaaatgaga aggtgctccc taagcactcg
     5821 ctcctgtacg agtacttcac agtctacaac gagctgacta aggtgaagta tgtgaccgag
     5881 ggcatgagga agccggcttt cctgtctggg gagcagaaga aggccatcgt ggacctcctg
     5941 ttcaagacca accggaaggt cacggttaag cagctcaagg aggactactt caagaagatt
     6001 gagtgcttcg attcggtcga gatctctggc gttgaggacc gcttcaacgc ctccctgggg
     6061 acctaccacg atctcctgaa gatcattaag gataaggact tcctggacaa cgaggagaat
     6121 gaggatatcc tcgaggacat tgtgctgaca ctcactctgt tcgaggaccg ggagatgatc
     6181 gaggagcgcc tgaagactta cgcccatctc ttcgatgaca aggtcatgaa gcagctcaag
     6241 aggaggaggt acaccggctg ggggaggctg agcaggaagc tcatcaacgg cattcgggac
     6301 aagcagtccg ggaagacgat cctcgacttc ctgaagagcg atggcttcgc gaaccgcaat
     6361 ttcatgcagc tgattcacga tgacagcctc acattcaagg aggatatcca gaaggctcag
     6421 gtgagcggcc agggggactc gctgcacgag catatcgcga acctcgctgg ctcgccagct
     6481 atcaagaagg ggattctgca gaccgtgaag gttgtggacg agctggtgaa ggtcatgggc
     6541 aggcacaagc ctgagaacat cgtcattgag atggcccggg agaatcagac cacgcagaag
     6601 ggccagaaga actcacgcga gaggatgaag aggatcgagg agggcattaa ggagctgggg
     6661 tcccagatcc tcaaggagca cccggtggag aacacgcagc tgcagaatga gaagctctac
     6721 ctgtactacc tccagaatgg ccgcgatatg tatgtggacc aggagctgga tattaacagg
     6781 ctcagcgatt acgacgtcga tcatatcgtt ccacagtcat tcctgaagga tgactccatt
     6841 gacaacaagg tcctcaccag gtcggacaag aaccggggca agtctgataa tgttccttca
     6901 gaggaggtcg ttaagaagat gaagaactac tggcgccagc tcctgaatgc caagctgatc
     6961 acgcagcgga agttcgataa cctcacaaag gctgagaggg gcgggctctc tgagctggac
     7021 aaggcgggct tcatcaagag gcagctggtc gagacacggc agatcactaa gcacgttgcg
     7081 cagattctcg actcacggat gaacactaag tacgatgaga atgacaagct gatccgcgag
     7141 gtgaaggtca tcaccctgaa gtcaaagctc gtctccgact tcaggaagga tttccagttc
     7201 tacaaggttc gggagatcaa caattaccac catgcccatg acgcgtacct gaacgcggtg
     7261 gtcggcacag ctctgatcaa gaagtaccca aagctcgaga gcgagttcgt gtacggggac
     7321 tacaaggttt acgatgtgag gaagatgatc gccaagtcgg agcaggagat tggcaaggct
     7381 accgccaagt acttcttcta ctctaacatt atgaatttct tcaagacaga gatcactctg
     7441 gccaatggcg agatccggaa gcgccccctc atcgagacga acggcgagac gggggagatc
     7501 gtgtgggaca agggcaggga tttcgcgacc gtcaggaagg ttctctccat gccacaagtg
     7561 aatatcgtca agaagacaga ggtccagact ggcgggttct ctaaggagtc aattctgcct
     7621 aagcggaaca gcgacaagct catcgcccgc aagaaggact gggatccgaa gaagtacggc
     7681 gggttcgaca gccccactgt ggcctactcg gtcctggttg tggcgaaggt tgagaagggc
     7741 aagtccaaga agctcaagag cgtgaaggag ctgctgggga tcacgattat ggagcgctcc
     7801 agcttcgaga agaacccgat cgatttcctg gaggcgaagg gctacaagga ggtgaagaag
     7861 gacctgatca ttaagctccc caagtactca ctcttcgagc tggagaacgg caggaagcgg
     7921 atgctggctt ccgctggcga gctgcagaag gggaacgagc tggctctgcc gtccaagtat
     7981 gtgaacttcc tctacctggc ctcccactac gagaagctca agggcagccc cgaggacaac
     8041 gagcagaagc agctgttcgt cgagcagcac aagcattacc tcgacgagat cattgagcag
     8101 atttccgagt tctccaagcg cgtgatcctg gccgacgcga atctggataa ggtcctctcc
     8161 gcgtacaaca agcaccgcga caagccaatc agggagcagg ctgagaatat cattcatctc
     8221 ttcaccctga cgaacctcgg cgcccctgct gctttcaagt acttcgacac aactatcgat
     8281 cgcaagaggt acacaagcac taaggaggtc ctggacgcga ccctcatcca ccagtcgatt
     8341 accggcctct acgagacgcg catcgacctg tctcagctcg ggggcgactc cggcggcagc
     8401 ccaaagaaga agcggaaggt gtctggaggt tctcctaaga aaaagagaaa agtgtccggc
     8461 ggctccccga agaagaagcg caaggtgtag gagctcagag ctttcgttcg tatcatcggt
     8521 ttcgacaacg ttcgtcaagt tcaatgcatc agtttcattg cgcacacacc agaatcctac
     8581 tgagtttgag tattatggca ttgggaaaac tgtttttctt gtaccatttg ttgtgcttgt
     8641 aatttactgt gttttttatt cggttttcgc tatcgaactg tgaaatggaa atggatggag
     8701 aagagttaat gaatgatatg gtccttttgt tcattctcaa attaatatta tttgtttttt
     8761 ctcttatttg ttgtgtgttg aatttgaaat tataagagat atgcaaacat tttgttttga
     8821 gtaaaaatgt gtcaaatcgt ggcctctaat gaccgaagtt aatatgagga gtaaaacact
     8881 tgtagttgta ccattatgct tattcactag gcaacaaata tattttcaga cctagaaaag
     8941 ctgcaaatgt tactgaatac aagtatgtcc tcttgtgttt tagacattta tgaactttcc
     9001 tttatgtaat tttccagaat ccttgtcaga ttctaatcat tgctttataa ttatagttat
     9061 actcatggat ttgtagttga gtatgaaaat attttttaat gcattttatg acttgccaat
     9121 tgattgacaa cgaattcgta atcatgtcat agctgtttcc tgtgtgaaat tgttatccgc
     9181 tcacaattcc acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat
     9241 gagtgagcta actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc
     9301 tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg
     9361 gctagagcag cttgccaaca tggtggagca cgacactctc gtctactcca agaatatcaa
     9421 agatacagtc tcagaagacc aaagggctat tgagactttt caacaaaggg taatatcggg
     9481 aaacctcctc ggattccatt gcccagctat ctgtcacttc atcaaaagga cagtagaaaa
     9541 ggaaggtggc acctacaaat gccatcattg cgataaagga aaggctatcg ttcaagatgc
     9601 ctctgccgac agtggtccca aagatggacc cccacccacg aggagcatcg tggaaaaaga
     9661 agacgttcca accacgtctt caaagcaagt ggattgatgt gaacatggtg gagcacgaca
     9721 ctctcgtcta ctccaagaat atcaaagata cagtctcaga agaccaaagg gctattgaga
     9781 cttttcaaca aagggtaata tcgggaaacc tcctcggatt ccattgccca gctatctgtc
     9841 acttcatcaa aaggacagta gaaaaggaag gtggcaccta caaatgccat cattgcgata
     9901 aaggaaaggc tatcgttcaa gatgcctctg ccgacagtgg tcccaaagat ggacccccac
     9961 ccacgaggag catcgtggaa aaagaagacg ttccaaccac gtcttcaaag caagtggatt
    10021 gatgtgatat ctccactgac gtaagggatg acgcacaatc ccactatcct tcgcaagacc
    10081 cttcctctat ataaggaagt tcatttcatt tggagaggac acgctgaaat caccagtctc
    10141 tctctacaaa tctatctctc tcgagctttc gcagatccgg ggggcaatga gatatgaaaa
    10201 agcctgaact caccgcgacg tctgtcgaga agtttctgat cgaaaagttc gacagcgtct
    10261 ccgacctgat gcagctctcg gagggcgaag aatctcgtgc tttcagcttc gatgtaggag
    10321 ggcgtggata tgtcctgcgg gtaaatagct gcgccgatgg tttctacaaa gatcgttatg
    10381 tttatcggca ctttgcatcg gccgcgctcc cgattccgga agtgcttgac attggggagt
    10441 ttagcgagag cctgacctat tgcatctccc gccgttcaca gggtgtcacg ttgcaagacc
    10501 tgcctgaaac cgaactgccc gctgttctac aaccggtcgc ggaggctatg gatgcgatcg
    10561 ctgcggccga tcttagccag acgagcgggt tcggcccatt cggaccgcaa ggaatcggtc
    10621 aatacactac atggcgtgat ttcatatgcg cgattgctga tccccatgtg tatcactggc
    10681 aaactgtgat ggacgacacc gtcagtgcgt ccgtcgcgca ggctctcgat gagctgatgc
    10741 tttgggccga ggactgcccc gaagtccggc acctcgtgca cgcggatttc ggctccaaca
    10801 atgtcctgac ggacaatggc cgcataacag cggtcattga ctggagcgag gcgatgttcg
    10861 gggattccca atacgaggtc gccaacatct tcttctggag gccgtggttg gcttgtatgg
    10921 agcagcagac gcgctacttc gagcggaggc atccggagct tgcaggatcg ccacgactcc
    10981 gggcgtatat gctccgcatt ggtcttgacc aactctatca gagcttggtt gacggcaatt
    11041 tcgatgatgc agcttgggcg cagggtcgat gcgacgcaat cgtccgatcc ggagccggga
    11101 ctgtcgggcg tacacaaatc gcccgcagaa gcgcggccgt ctggaccgat ggctgtgtag
    11161 aagtactcgc cgatagtgga aaccgacgcc ccagcactcg tccgagggca aagaaataga
    11221 gtagatgccg accgggatct gtcgatcgac aagctcgagt ttctccataa taatgtgtga
    11281 gtagttccca gataagggaa ttagggttcc tatagggttt cgctcatgtg ttgagcatat
    11341 aagaaaccct tagtatgtat ttgtatttgt aaaatacttc tatcaataaa atttctaatt
    11401 cctaaaacca aaatccagta ctaaaatcca gatcccccga attaattcgg cgttaattca
    11461 gtacattaaa aacgtccgca atgtgttatt aagttgtcta agcgtcaatt tgtttacacc
    11521 acaatatatc ctgccaccag ccagccaaca gctccccgac cggcagctcg gcacaaaatc
    11581 accactcgat acaggcagcc catcagtccg ggacggcgtc agcgggagag ccgttgtaag
    11641 gcggcagact ttgctcatgt taccgatgct attcggaaga acggcaacta agctgccggg
    11701 tttgaaacac ggatgatctc gcggagggta gcatgttgat tgtaacgatg acagagcgtt
    11761 gctgcctgtg atcaccgcgg tttcaaaatc ggctccgtcg atactatgtt atacgccaac
    11821 tttgaaaaca actttgaaaa agctgttttc tggtatttaa ggttttagaa tgcaaggaac
    11881 agtgaattgg agttcgtctt gttataatta gcttcttggg gtatctttaa atactgtaga
    11941 aaagaggaag gaaataataa atggctaaaa tgagaatatc accggaattg aaaaaactga
    12001 tcgaaaaata ccgctgcgta aaagatacgg aaggaatgtc tcctgctaag gtatataagc
    12061 tggtgggaga aaatgaaaac ctatatttaa aaatgacgga cagccggtat aaagggacca
    12121 cctatgatgt ggaacgggaa aaggacatga tgctatggct ggaaggaaag ctgcctgttc
    12181 caaaggtcct gcactttgaa cggcatgatg gctggagcaa tctgctcatg agtgaggccg
    12241 atggcgtcct ttgctcggaa gagtatgaag atgaacaaag ccctgaaaag attatcgagc
    12301 tgtatgcgga gtgcatcagg ctctttcact ccatcgacat atcggattgt ccctatacga
    12361 atagcttaga cagccgctta gccgaattgg attacttact gaataacgat ctggccgatg
    12421 tggattgcga aaactgggaa gaagacactc catttaaaga tccgcgcgag ctgtatgatt
    12481 ttttaaagac ggaaaagccc gaagaggaac ttgtcttttc ccacggcgac ctgggagaca
    12541 gcaacatctt tgtgaaagat ggcaaagtaa gtggctttat tgatcttggg agaagcggca
    12601 gggcggacaa gtggtatgac attgccttct gcgtccggtc gatcagggag gatatcgggg
    12661 aagaacagta tgtcgagcta ttttttgact tactggggat caagcctgat tgggagaaaa
    12721 taaaatatta tattttactg gatgaattgt tttagtacct agaatgcatg accaaaatcc
    12781 cttaacgctg atgaatcccc taatgatttt ggtaaaaatc attaagttaa ggtggataca
    12841 catcttgtca tatgatcaaa tggtttcgcg aaaaatcaat aatcagacaa caagatgtgc
    12901 gaactcgata ttttacacga ctctctttac caattctgcc ccgaattaca cttaaaacga
    12961 ctcaacagct taacgttggc ttgccacgca ttacttgact gtaaaactct cactcttacc
    13021 gaacttggcc gtaacctgcc aaccaaagcg agaacaaaac ataacatcaa acgaatcgac
    13081 cgattgttag gtaatcgtca cctccacaaa gagcgactcg ctgtataccg ttggcatgct
    13141 agctttatct gttcgggcaa tacgatgccc attgtacttg ttgactggtc tgatattcgt
    13201 gagcaaaaac gacttatggt attgcgagct tcagtcgcac tacacggtcg ttctgttact
    13261 ctttatgaga aagcgttccc gctttcagag caatgttcaa agaaagctca tgaccaattt
    13321 ctagccgacc ttgcgagcat tctaccgagt aacaccacac cgctcattgt cagtgatgct
    13381 ggctttaaag tgccatggta taaatccgtt gagaagctgg gttggtactg gttaagtcga
    13441 gtaagaggaa aagtacaata tgcagaccta ggagcggaaa actggaaacc tatcagcaac
    13501 ttacatgata tgtcatctag tcactcaaag actttaggct ataagaggct gactaaaagc
    13561 aatccaatct catgccaaat tctattgtat aaatctcgct ctaaaggccg aaaaaatcag
    13621 cgctcgacac ggactcattg tcaccacccg tcacctaaaa tctactcagc gtcggcaaag
    13681 gagccatggg ttctagcaac taacttacct gttgaaattc gaacacccaa acaacttgtt
    13741 aatatctatt cgaagcgaat gcagattgaa gaaaccttcc gagacttgaa aagtcctgcc
    13801 tacggactag gcctacgcca tagccgaacg agcagctcag agcgttttga tatcatgctg
    13861 ctaatcgccc tgatgcttca actaacatgt tggcttgcgg gcgttcatgc tcagaaacaa
    13921 ggttgggaca agcacttcca ggctaacaca gtcagaaatc gaaacgtact ctcaacagtt
    13981 cgcttaggca tggaagtttt gcggcattct ggctacacaa taacaaggga agacttactc
    14041 gtggctgcaa ccctactagc tcaaaattta ttcacacatg gttacgcttt ggggaaatta
    14101 tgaggggatc tctcagccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
    14161 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
    14221 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
    14281 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc
    14341 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
    14401 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
    14461 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
    14521 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
    14581 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
    14641 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
    14701 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
    14761 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
    14821 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
    14881 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
    14941 aagcggaaga gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
    15001 gcatatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagtatacac
    15061 tccgctatcg ctacgtgact gggtcatggc tgcgccccga cacccgccaa cacccgctga
    15121 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
    15181 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga ggcagggtgc
    15241 cttgatgtgg gcgccggcgg tcgagtggcg acggcgcggc ttgtccgcgc cctggtagat
    15301 tgcctggccg taggccagcc atttttgagc ggccagcggc cgcgataggc cgacgcgaag
    15361 cggcggggcg tagggagcgc agcgaccgaa gggtaggcgc tttttgcagc tcttcggctg
    15421 tgcgctggcc agacagttat gcacaggcca ggcgggtttt aagagtttta ataagtttta
    15481 aagagtttta ggcggaaaaa tcgccttttt tctcttttat atcagtcact tacatgtgtg
    15541 accggttccc aatgtacggc tttgggttcc caatgtacgg gttccggttc ccaatgtacg
    15601 gctttgggtt cccaatgtac gtgctatcca caggaaacag accttttcga cctttttccc
    15661 ctgctagggc aatttgccct agcatctgct ccgtacatta ggaaccggcg gatgcttcgc
    15721 cctcgatcag gttgcggtag cgcatgacta ggatcgggcc agcctgcccc gcctcctcct
    15781 tcaaatcgta ctccggcagg tcatttgacc cgatcagctt gcgcacggtg aaacagaact
    15841 tcttgaactc tccggcgctg ccactgcgtt cgtagatcgt cttgaacaac catctggctt
    15901 ctgccttgcc tgcggcgcgg cgtgccaggc ggtagagaaa acggccgatg ccgggatcga
    15961 tcaaaaagta atcggggtga accgtcagca cgtccgggtt cttgccttct gtgatctcgc
    16021 ggtacatcca atcagctagc tcgatctcga tgtactccgg ccgcccggtt tcgctcttta
    16081 cgatcttgta gcggctaatc aaggcttcac cctcggatac cgtcaccagg cggccgttct
    16141 tggccttctt cgtacgctgc atggcaacgt gcgtggtgtt taaccgaatg caggtttcta
    16201 ccaggtcgtc tttctgcttt ccgccatcgg ctcgccggca gaacttgagt acgtccgcaa
    16261 cgtgtggacg gaacacgcgg ccgggcttgt ctcccttccc ttcccggtat cggttcatgg
    16321 attcggttag atgggaaacc gccatcagta ccaggtcgta atcccacaca ctggccatgc
    16381 cggccggccc tgcggaaacc tctacgtgcc cgtctggaag ctcgtagcgg atcacctcgc
    16441 cagctcgtcg gtcacgcttc gacagacgga aaacggccac gtccatgatg ctgcgactat
    16501 cgcgggtgcc cacgtcatag agcatcggaa cgaaaaaatc tggttgctcg tcgcccttgg
    16561 gcggcttcct aatcgacggc gcaccggctg ccggcggttg ccgggattct ttgcggattc
    16621 gatcagcggc cgcttgccac gattcaccgg ggcgtgcttc tgcctcgatg cgttgccgct
    16681 gggcggcctg cgcggccttc aacttctcca ccaggtcatc acccagcgcc gcgccgattt
    16741 gtaccgggcc ggatggtttg cgaccgctca cgccgattcc tcgggcttgg gggttccagt
    16801 gccattgcag ggccggcagg caacccagcc gcttacgcct ggccaaccgc ccgttcctcc
    16861 acacatgggg cattccacgg cgtcggtgcc tggttgttct tgattttcca tgccgcctcc
    16921 tttagccgct aaaattcatc tactcattta ttcatttgct catttactct ggtagctgcg
    16981 cgatgtattc agatagcagc tcggtaatgg tcttgccttg gcgtaccgcg tacatcttca
    17041 gcttggtgtg atcctccgcc ggcaactgaa agttgacccg cttcatggct ggcgtgtctg
    17101 ccaggctggc caacgttgca gccttgctgc tgcgtgcgct cggacggccg gcacttagcg
    17161 tgtttgtgct tttgctcatt ttctctttac ctcattaact caaatgagtt ttgatttaat
    17221 ttcagcggcc agcgcctgga cctcgcgggc agcgtcgccc tcgggttctg attcaagaac
    17281 ggttgtgccg gcggcggcag tgcctgggta gctcacgcgc tgcgtgatac gggactcaag
    17341 aatgggcagc tcgtacccgg ccagcgcctc ggcaacctca ccgccgatgc gcgtgccttt
    17401 gatcgcccgc gacacgacaa aggccgcttg tagccttcca tccgtgacct caatgcgctg
    17461 cttaaccagc tccaccaggt cggcggtggc ccatatgtcg taagggcttg gctgcaccgg
    17521 aatcagcacg aagtcggctg ccttgatcgc ggacacagcc aagtccgccg cctggggcgc
    17581 tccgtcgatc actacgaagt cgcgccggcc gatggccttc acgtcgcggt caatcgtcgg
    17641 gcggtcgatg ccgacaacgg ttagcggttg atcttcccgc acggccgccc aatcgcgggc
    17701 actgccctgg ggatcggaat cgactaacag aacatcggcc ccggcgagtt gcagggcgcg
    17761 ggctagatgg gttgcgatgg tcgtcttgcc tgacccgcct ttctggttaa gtacagcgat
    17821 aaccttcatg cgttcccctt gcgtatttgt ttatttactc atcgcatcat atacgcagcg
    17881 accgcatgac gcaagctgtt ttactcaaat acacatcacc tttttagacg gcggcgctcg
    17941 gtttcttcag cggccaagct ggccggccag gccgccagct tggcatcaga caaaccggcc
    18001 aggatttcat gcagccgcac ggttgagacg tgcgcgggcg gctcgaacac gtacccggcc
    18061 gcgatcatct ccgcctcgat ctcttcggta atgaaaaacg gttcgtcctg gccgtcctgg
    18121 tgcggtttca tgcttgttcc tcttggcgtt cattctcggc ggccgccagg gcgtcggcct
    18181 cggtcaatgc gtcctcacgg aaggcaccgc gccgcctggc ctcggtgggc gtcacttcct
    18241 cgctgcgctc aagtgcgcgg tacagggtcg agcgatgcac gccaagcagt gcagccgcct
    18301 ctttcacggt gcggccttcc tggtcgatca gctcgcgggc gtgcgcgatc tgtgccgggg
    18361 tgagggtagg gcgggggcca aacttcacgc ctcgggcctt ggcggcctcg cgcccgctcc
    18421 gggtgcggtc gatgattagg gaacgctcga actcggcaat gccggcgaac acggtcaaca
    18481 ccatgcggcc ggccggcgtg gtggtgtcgg cccacggctc tgccaggcta cgcaggcccg
    18541 cgccggcctc ctggatgcgc tcggcaatgt ccagtaggtc gcgggtgctg cgggccaggc
    18601 ggtctagcct ggtcactgtc acaacgtcgc cagggcgtag gtggtcaagc atcctggcca
    18661 gctccgggcg gtcgcgcctg gtgccggtga tcttctcgga aaacagcttg gtgcagccgg
    18721 ccgcgtgcag ttcggcccgt tggttggtca agtcctggtc gtcggtgctg acgcgggcat
    18781 agcccagcag gccagcggcg gcgctcttgt tcatggcgta atgtctccgg ttctagtcgc
    18841 aagtattcta ctttatgcga ctaaaacac
//

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