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  • pCas-Guide-Nickase

    價格:2000元

    聯系方式:I47-825O-882O

    買家導航

    載體基本信息

    出品公司: OriGene
    載體名稱: pCas-Guide-Nickase
    質粒類型: CRISPR/Cas9載體;基因敲除載體;哺乳動物細胞載體
    高拷貝/低拷貝: 高拷貝
    插入位點: 限制性內切酶,多克隆位點
    啟動子: U6
    載體大小: 8033 bp
    5' 測序引物及序列: 5’ gatcgATGGGAGGTGGTATGGGAGGg 3’
    3' 測序引物及序列: 5’ aaaacCCTCCCATACCACCTCCCATc 3’
    載體標簽: Myc(C-ter)
    載體抗性: 氨芐青霉素
    篩選標記: --
    克隆菌株: Stbl3
    宿主細胞(系): 哺乳動物細胞
    備注: --
    產品目錄號: GE100019
    穩定性: 瞬表達
    組成型/誘導型: 組成型
    病毒/非病毒: 非病毒

    載體質粒圖譜和多克隆位點信息

    pCas-Guide-Nickase載體圖譜



    載體簡介

    pCas-Guide-Nickase載體特征
    
    Expresses human codon-optimized Cas9 nickase D10A mutant, C-terminal Myc-DDK tagged
    BamH I and BsmB I can be used for genomic target sequence cloning
    Expresses gRNA containing the inserted target sequence and the gRNA scaffold
    CF3 primer can be used to sequence the target sequence after cloning
    

    載體序列

    LOCUS       Exported File           8033 bp ds-DNA    circular SYN 23-七月-2016
    DEFINITION  Mammalian vector for co-expressing a guide RNA (gRNA) together with 
                the D10A nickase mutant of Cas9, for genome editing using the 
                CRISPR/Cas9 system.
    ACCESSION   .
    VERSION     .
    KEYWORDS    pCas-Guide-Nickase
    SOURCE      synthetic DNA construct
      ORGANISM  synthetic DNA construct
    REFERENCE   1  (bases 1 to 8033)
      AUTHORS   OriGene
      TITLE     Direct Submission
      JOURNAL   Exported 2016-07-23 from SnapGene Viewer 1.5.3
                http://www.snapgene.com
    FEATURES             Location/Qualifiers
         source          1..8033
                         /organism="synthetic DNA construct"
                         /lab_host="Mammalian Cells"
                         /mol_type="other DNA"
         promoter        90..330
                         /note="U6 promoter"
                         /note="RNA polymerase III promoter for human U6 snRNA"
                         /note="color: #000000; direction: RIGHT"
         misc_RNA        384..459
                         /note="gRNA scaffold"
                         /note="guide RNA scaffold for the Streptococcus pyogenes 
                         CRISPR/Cas9 system"
                         /note="color: #00ccff; direction: RIGHT"
         terminator      460..465
                         /note="polIII terminator"
                         /note="RNA polymerase III transcription terminator"
                         /note="color: #ffffff"
         enhancer        511..890
                         /note="CMV enhancer"
                         /note="human cytomegalovirus immediate early enhancer"
                         /note="color: #000000"
         promoter        891..1094
                         /note="CMV promoter"
                         /note="human cytomegalovirus (CMV) immediate early 
                         promoter"
                         /note="color: #000000; direction: RIGHT"
         promoter        1120..1138
                         /note="T7 promoter"
                         /note="promoter for bacteriophage T7 RNA polymerase"
                         /note="color: #ffffff; direction: RIGHT"
         CDS             1197..5300
                         /codon_start=1
                         /locus_tag="
                         "
                         /product="nickase mutant of the Cas9 endonuclease from 
                         the?Streptococcus pyogenes?Type II CRISPR/Cas system"
                         /note="Cas9(D10A)"
                         /note="generates RNA-guided single strand nicks in DNA due 
                         to the D10A mutation in the RuvC catalytic domain"
                         /note="This feature has 3 segments:
                         ???1:?1197?..?1223?/?#993366
                         ???2:?1224?..?1226?/?#cc99b2
                         ???3:?1227?..?5300?/?#993366"
                         /protein_id="
                         "
                         /translation="MDKKYSIGLAIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                         NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                         FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                         FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                         ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                         IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                         QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                         RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                         NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                         FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                         SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                         LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                         QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                         HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                         LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                         SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                         VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                         NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                         EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                         ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                         TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                         LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                         ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                         DATLIHQSITGLYETRIDLSQLGGD"
         CDS             5301..5321
                         /codon_start=1
                         /locus_tag="
                         "
                         /product="nuclear localization signal of SV40 large T 
                         antigen"
                         /note="SV40 NLS"
                         /note="
                         "
                         /note="color: #993366"
                         /protein_id="
                         "
                         /translation="PKKKRKV"
         CDS             5328..5348
                         /codon_start=1
                         /locus_tag="
                         "
                         /product="nuclear localization signal of SV40 large T 
                         antigen"
                         /note="SV40 NLS"
                         /note="
                         "
                         /note="color: #993366"
                         /protein_id="
                         "
                         /translation="PKKKRKV"
         CDS             5370..5399
                         /codon_start=1
                         /product="Myc (human c-Myc oncogene) epitope tag"
                         /note="Myc"
                         /note="color: #cc99b2"
                         /translation="EQKLISEEDL"
         CDS             5418..5441
                         /codon_start=1
                         /product="FLAG(R) epitope tag, followed by an enterokinase 
                         cleavage site"
                         /note="FLAG"
                         /note="color: #cc99b2"
                         /translation="DYKDDDDK"
         polyA_signal    5488..6110
                         /note="hGH poly(A) signal"
                         /note="human growth hormone polyadenylation signal"
                         /note="color: #a6acb3"
         rep_origin      complement(6231..6819)
                         /direction=LEFT
                         /note="ori"
                         /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                         replication"
                         /note="color: #ffff00"
         CDS             complement(6994..7854)
                         /codon_start=1
                         /gene="bla"
                         /product="beta-lactamase"
                         /note="AmpR"
                         /note="confers resistance to ampicillin, carbenicillin, and
                         related antibiotics"
                         /note="This feature has 2 segments:
                         ???1:?6994?..?7785?/?#ccffcc
                         ???2:?7786?..?7854?/?#ccffcc?/?signal?sequence
                         Cleavage site after base 7785"
                         /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                         ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                         PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                         EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                         LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                         LIKHW"
    ORIGIN
            1 gaattcccca gtggaaagac gcgcaggcaa aacgcaccac gtgacggagc gtgaccgcgc
           61 gccgagcgcg cgccaaggtc gggcaggaag agggcctatt tcccatgatt ccttcatatt
          121 tgcatatacg atacaaggct gttagagaga taattagaat taatttgact gtaaacacaa
          181 agatattagt acaaaatacg tgacgtagaa agtaataatt tcttgggtag tttgcagttt
          241 taaaattatg ttttaaaatg gactatcata tgcttaccgt aacttgaaag tatttcgatt
          301 tcttgggttt atatatcttg tggaaaggac gcgggatcca ctggaccagg cagcagcgtc
          361 agaagacttt tttggaacgt ctcgttttag agctagaaat agcaagttaa aataaggcta
          421 gtccgttatc aacttgaaaa agtggcaccg agtcggtgct ttttttggtg tacatttata
          481 ttggctcatg tccaatatga ccgccatgtt gacattgatt attgactagt tattaatagt
          541 aatcaattac ggggtcatta gttcatagcc catatatgga gttccgcgtt acataactta
          601 cggtaaatgg cccgcctggc tgaccgccca acgacccccg cccattgacg tcaataatga
          661 cgtatgttcc catagtaacg ccaataggga ctttccattg acgtcaatgg gtggagtatt
          721 tacggtaaac tgcccacttg gcagtacatc aagtgtatca tatgccaagt ccgcccccta
          781 ttgacgtcaa tgacggtaaa tggcccgcct ggcattatgc ccagtacatg accttacggg
          841 actttcctac ttggcagtac atctacgtat tagtcatcgc tattaccatg gtgatgcggt
          901 tttggcagta caccaatggg cgtggatagc ggtttgactc acggggattt ccaagtctcc
          961 accccattga cgtcaatggg agtttgtttt ggcaccaaaa tcaacgggac tttccaaaat
         1021 gtcgtaataa ccccgccccg ttgacgcaaa tgggcggtag gcgtgtacgg tgggaggtct
         1081 atataagcag agctcgttta gtgaaccgtc agaattttgt aatacgactc actatagggc
         1141 ggccgggaat tcgtcgactg gaaccggtac cgaggagatc tgccgccgcg atcgccatgg
         1201 ataagaaata ctcaatagga ctggctattg gcacaaatag cgtcggatgg gctgtgatca
         1261 ctgatgaata taaggttcct tctaaaaagt tcaaggttct gggaaataca gaccgccaca
         1321 gtatcaaaaa aaatcttata ggggctcttc tgtttgacag tggagagaca gccgaagcta
         1381 ctagactcaa acggacagct aggagaaggt atacaagacg gaagaatagg atttgttatc
         1441 tccaggagat tttttcaaat gagatggcca aagtggatga tagtttcttt catagacttg
         1501 aagagtcttt tttggtggaa gaagacaaga agcatgaaag acatcctatt tttggaaata
         1561 tagtggatga agttgcttat cacgagaaat atccaactat ctatcatctg agaaaaaaat
         1621 tggtggattc tactgataaa gccgatttgc gcctgatcta tttggccctg gcccacatga
         1681 ttaagtttag aggtcatttt ttgattgagg gcgatctgaa tcctgataat agtgatgtgg
         1741 acaaactgtt tatccagttg gtgcaaacct acaatcaact gtttgaagaa aaccctatta
         1801 acgcaagtgg agtggatgct aaagccattc tttctgcaag attgagtaaa tcaagaagac
         1861 tggaaaatct cattgctcag ctccccggtg agaagaaaaa tggcctgttt gggaatctca
         1921 ttgctttgtc attgggtttg acccctaatt ttaaatcaaa ttttgatttg gcagaagatg
         1981 ctaaactcca gctttcaaaa gatacttacg atgatgatct ggataatctg ttggctcaaa
         2041 ttggagatca atatgctgat ttgtttttgg cagctaagaa tctgtcagat gctattctgc
         2101 tttcagacat cctgagagtg aatactgaaa taactaaggc tcccctgtca gcttcaatga
         2161 ttaaacgcta cgatgaacat catcaagact tgactcttct gaaagccctg gttagacaac
         2221 aacttccaga aaagtataaa gaaatctttt ttgatcaatc aaaaaacgga tatgcaggtt
         2281 atattgatgg cggcgcaagc caagaagaat tttataaatt tatcaaacca attctggaaa
         2341 aaatggatgg tactgaggaa ctgttggtga aactgaatag agaagatttg ctgcgcaagc
         2401 aacggacctt tgacaacggc tctattcccc atcaaattca cttgggtgag ctgcatgcta
         2461 ttttgagaag acaagaagac ttttatccat ttctgaaaga caatagagag aagattgaaa
         2521 aaatcttgac ttttaggatt ccttattatg ttggtccatt ggccagaggc aatagtaggt
         2581 ttgcatggat gactcggaag tctgaagaaa caattacccc atggaatttt gaagaagttg
         2641 tcgataaagg tgcttcagct caatcattta ttgaacgcat gacaaacttt gataaaaatc
         2701 ttccaaatga aaaagtgctg ccaaaacata gtttgcttta tgagtatttt accgtttata
         2761 acgaattgac aaaggtcaaa tatgttactg aaggaatgag aaaaccagca tttctttcag
         2821 gtgaacagaa gaaagccatt gttgatctgc tcttcaaaac aaataggaaa gtgaccgtta
         2881 agcaactgaa agaagattat ttcaaaaaaa tagaatgttt tgatagtgtt gaaatttcag
         2941 gagttgaaga tagatttaat gcttcactgg gtacatacca tgatttgctg aaaattatta
         3001 aagataaaga ttttttggat aatgaagaaa atgaagacat cctggaggat attgttctga
         3061 cattgaccct gtttgaagat agggagatga ttgaggaaag acttaaaaca tacgctcacc
         3121 tctttgatga taaggtgatg aaacagctta aaagacgcag atatactggt tggggaaggt
         3181 tgtccagaaa attgattaat ggtattaggg ataagcaatc tggcaaaaca atactggatt
         3241 ttttgaaatc agatggtttt gccaatcgca attttatgca gctcatccat gatgatagtt
         3301 tgacatttaa agaagacatc caaaaagcac aagtgtctgg acaaggcgat agtctgcatg
         3361 aacatattgc aaatctggct ggtagccctg ctattaaaaa aggtattctc cagactgtga
         3421 aagttgttga tgaattggtc aaagtgatgg ggcggcataa gccagaaaat atcgttattg
         3481 aaatggcaag agaaaatcag acaactcaaa agggccagaa aaattccaga gagaggatga
         3541 aaagaatcga agaaggtatc aaagaactgg gaagtcagat tcttaaagag catcctgttg
         3601 aaaatactca attgcaaaat gaaaagctct atctctatta tctccaaaat ggaagagata
         3661 tgtatgtgga ccaagaactg gatattaata ggctgagtga ttatgatgtc gatcacattg
         3721 ttccacaaag tttccttaaa gacgattcaa tagacaataa ggtcctgacc aggtctgata
         3781 aaaatagagg taaatccgat aacgttccaa gtgaagaagt ggtcaaaaag atgaaaaact
         3841 attggagaca acttctgaac gccaagctga tcactcaaag gaagtttgat aatctgacca
         3901 aagctgaaag aggaggtttg agtgaacttg ataaagctgg ttttatcaaa cgccaattgg
         3961 ttgaaactcg ccaaatcact aagcatgtgg cacaaatttt ggatagtcgc atgaatacta
         4021 aatacgatga aaatgataaa cttattagag aggttaaagt gattaccctg aaatctaaac
         4081 tggtttctga cttcagaaaa gatttccaat tctataaagt gagagagatt aacaattacc
         4141 atcatgccca tgatgcctat ctgaatgccg tcgttggaac tgctttgatt aagaaatatc
         4201 caaaacttga aagcgagttt gtctatggtg attataaagt ttatgatgtt aggaaaatga
         4261 ttgctaagtc tgagcaagaa ataggcaaag caaccgcaaa gtatttcttt tactctaata
         4321 tcatgaactt cttcaaaaca gaaattacac ttgcaaatgg agagattcgc aaacgccctc
         4381 tgatcgaaac taatggggaa actggagaaa ttgtctggga taaagggaga gattttgcca
         4441 cagtgcgcaa agtgttgtcc atgccccaag tcaatatcgt caagaaaaca gaagtgcaga
         4501 caggcggatt ctctaaggag tcaattctgc caaaaagaaa ttccgacaag ctgattgcta
         4561 ggaaaaaaga ctgggaccca aaaaaatatg gtggttttga tagtccaacc gtggcttatt
         4621 cagtcctggt ggttgctaag gtggaaaaag ggaaatccaa gaagctgaaa tccgttaaag
         4681 agctgctggg gatcacaatt atggaaagaa gttcctttga aaaaaatccc attgactttc
         4741 tggaagctaa aggatataag gaagttaaaa aagacctgat cattaaactg cctaaatata
         4801 gtctttttga gctggaaaac ggtaggaaac ggatgctggc tagtgccgga gaactgcaaa
         4861 aaggaaatga gctggctctg ccaagcaaat atgtgaattt tctgtatctg gctagtcatt
         4921 atgaaaagtt gaagggtagt ccagaagata acgaacaaaa acaattgttt gtggagcagc
         4981 ataagcatta tctggatgag attattgagc aaatcagtga attttctaag agagttattc
         5041 tggcagatgc caatctggat aaagttctta gtgcatataa caaacataga gacaaaccaa
         5101 taagagaaca agcagaaaat atcattcatc tgtttacctt gaccaatctt ggagcacccg
         5161 ctgcttttaa atactttgat acaacaattg ataggaaaag atatacctct acaaaagaag
         5221 ttctggatgc cactcttatc catcaatcca tcactggtct ttatgaaaca cgcattgatt
         5281 tgagtcagct gggaggtgac cccaagaaaa aacgcaaggt ggaagatcct aagaaaaagc
         5341 ggaaagtgga cacgcgtacg cggccgctcg agcagaaact catctcagaa gaggatctgg
         5401 cagcaaatga tatcctggat tacaaggatg acgacgataa ggtttaaacg gccggccgcg
         5461 gtcatagctg tttcctgaac agatcccggg tggcatccct gtgacccctc cccagtgcct
         5521 ctcctggccc tggaagttgc cactccagtg cccaccagcc ttgtcctaat aaaattaagt
         5581 tgcatcattt tgtctgacta ggtgtccttc tataatatta tggggtggag gggggtggta
         5641 tggagcaagg ggcaagttgg gaagacaacc tgtagggcct gcggggtcta ttgggaacca
         5701 agctggagtg cagtggcaca atcttggctc actgcaatct ccgcctcctg ggttcaagcg
         5761 attctcctgc ctcagcctcc cgagttgttg ggattccagg catgcatgac caggctcagc
         5821 taatttttgt ttttttggta gaggcggggt ttcaccatat tggccaggct ggtctccaac
         5881 tcctaatctc aggtgatcta cccaccttgg cctcccaaat tgctgggatt acaggcgtga
         5941 accactgctc ccttccctgt ccttctgatt ttaaaataac tataccagca ggaggacgtc
         6001 cagacacagc ataggctacc tggccatgcc caaccggtgg gacatttgag ttgcttgctt
         6061 ggcactgtcc tctcatgcgt tgggtccact cagtagatgc ctgttgaatt gggtacgcgg
         6121 ccagcggcga gcggtatcag ctcactcaaa ggcggtaata cggttatcca cagaatcagg
         6181 ggataacgca ggaaagaaca tgtccgtaaa aaggccgcgt tgctggcgtt tttccatagg
         6241 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
         6301 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
         6361 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
         6421 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
         6481 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
         6541 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
         6601 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
         6661 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
         6721 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
         6781 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
         6841 acggggtctg acgctcagtg gaacgacgcg taactcacgt taagggattt tggtcatgag
         6901 attatcaaaa aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat
         6961 ctaaagtata tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc
         7021 tatctcagcg atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat
         7081 aactacgata cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc
         7141 acgctcaccg gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag
         7201 aagtggtcct gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag
         7261 agtaagtagt tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt
         7321 ggtgtcacgc tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg
         7381 agttacatga tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt
         7441 tgtcagaagt aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc
         7501 tcttactgtc atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc
         7561 attctgagaa tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa
         7621 taccgcgcca catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg
         7681 aaaactctca aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc
         7741 caactgatct tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag
         7801 gcaaaatgcc gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt
         7861 cctttttcaa tattattgaa gcatttatca gggttattgt ctcatgatga tattatttta
         7921 tcttgtgcaa tgtaacatca gagattttga gacacgggcc agagctgcca ggaaacagct
         7981 atgaccatgt aatacgactc actatagggg atatcagctg gatggcagtt aac
    //
    
    

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