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  • weird 454 reads

    Hello
    I have a weird 454 result produced by MID and DSN normalization.
    The mRNA reads started with 'key(4bp)+MID(10bp)+cDNA library primer (20~25bp) sequence from 5' end. The reads are supposed to included key+MID, not cDNA library primer. I feel like that there seemed to be something wrong with cDNA library construction. Below is the sample reads.
    Are they normal?


    >GRMETAB01A1BYK
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTGGGAGAATCTCGAACCACATCTCTGCCCGTACCTAGCaacctaactctggtcgtnaagagctgntcncctcccncctcctntgtgtgtactactacgctcttntagacagcacgacgacacacggaggtaggagnn
    >GRMETAB01AK1L7
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTGACTGAAACACAATTCATTTCTGACTGACATTacctttctgtcaactagacggtggcaccacaaataataccggtctttgtgcttgtctcttctagatgganggtttgttaaggactcttgttgtcattactttaaaaagaagtaataaaggattttaaagtaacgaaaaaacgaaaagaaaaactaaaactaaacgtcgtgtagtacactacacgtctctgagagcacgacgacacacggaggtaggngnnnnn
    >GRMETAB01BC7E6
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTACGGAGGTGTTGAAGaagattgaatcaagaactaacagccaaagaagaagaagaagaagaagacagaagaggagcagaagaaactaagacacagattcaagaatctcaggtaacatccaccaacaaggaacggacggttcctaacgacgggagtgagaactaagacaagaaatactcaagaccactggaggaggctaaagacagagcgagaaaacgaaagaacagaacaagctgagctgagactgccaacggacacacagggagtaggnnnn
    >GRMETAB01EOICN
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTACTGAGGGAAGTAGTTGTGTTGGCTGAGTTGTCAGTCAAGCGAATTCCGCTACTCTGCGTTGATACCACTGCTctgagactgccaaggcacacagggataggnnnnnnnnnn
    >GRMETAB01DR31L
    tcagACGAGTGCGTAGCAGTGGTATCAACGCAGAGTACTTTTTTTTTNTTTNTTTAttataactaactaactatatatattattattatatatatctctactactactactactatatactactatatatatactactactacaccacacacgtagtagtagtacgtactactactactacgagagagagagagagacgagtaggtagtacgtacgtacgtacgacgacgtactactaactaactaactacgtacgacgacgacgacgacgacgacgtacgtacgtaccgtaccgtaccgtaccgtaccgtaccgacctacctaacctacctaacctactactactacgtacgacgacgacgacgacgacgacgagagagngnnnn
    >GRMETAB01A4CCL
    tcagACGAGTGCGTAGCAGTGGTATCAACGCAGAGTTACTTTTTTTTTTTTTTTTTTTTTNttttnttttnttttnttttnttttntttatttatttatttatttatttatttattattattatatatatatatatatatatatatatatatatatatagtatatatatatatatatatatatatatatatatatatatatagtatagtagtagtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgacgacgacgacgacgacgagagagagagngnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn
    >GRMETAB01BL4UE
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTACTTTNTTTNTTTCTTTCTTTCTTTCTTTCttctctactatctacaanagacacacactctctctcgtctcgtcgtacgtacgtacgtcgacgacgacgacgacgacgacgagagagngnnn
    >GRMETAB01BWBZ9
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGACAGCCAAGTCATTGCCAGACACATACAAAGATAAGCCCACGTActtacngcgttgataccacgtcggcttctgaggacgttggaccaaagggacgacctaacctagggggaattaggggngcncncnngagagaggngcngtnttnanntgngnngagangcacagatacgctcgcagacgtgggttcggtccgacgacccgtaacggtaacctacaacgcttcctttaggttatagtaattaaatttagtacttatttcgttcggacggacgaacgaaaaacgggaaaccgtaacgtaccaccacgtacctactaggacaaacaactacttcttcgtcgtctagtaggacgagggaggacgtacgtaggtagtttattattaattacttctagaagacacagagacgacgaccaacgtactattcgaccgaacgaacaacaactaaacaaaacgaaacgtaaaccgacccgtaacgtacttacgtcgtcgttacgtaactaacctactcgtcttcgtacgacgacgtacgaccgaacggacgacgaacggaagggaaggtaggagnnnnn
    >GRMETAB01C584Q
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTACAGGGGTGTTTACAAGAGATGGTTTGCAATGCAGACTaagcagaaaatttagtagngtgatggataggaaaccaaaccaacagcccccgtacctctncgtgtatacacctgctcttcgagactgcacggacgacacaggggagtagngnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn
    >GRMETAB01AOKGH
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTACGTTTNTTTNTTTTTTTTTNTTTTTTNTTTTTNttttnttttntttttattttattttattttattttattttattttatttatttatttatttatttatttatttatttattacttacttattagtagtagtagtagtacgtagtacgtacgtacgtacgtacgtacgtacgtacgtactacgtacgtacgtacgtacgtacgtactactactactactactactactacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgtacgacgacgacgacgacgacgacgagacgacgagagannnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn
    >GRMETAB01D7O01
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTACGGGAATTATGACATGCAccgatttacatcccncctctcngtcgtatanacctatctccttcagaagtnttctacgagaaccaacctaggnggacgagatntctcntacgtcttgtacganctacctcgngnnntcncntctcgttgtggtggttcttacgtcgttcgttgtagttgtaagtaacgtacgtgagagaagacgtacgtacgactacctacgtacgtacaactaaagttaaagttaaattaaaaagtaaaaagttaaagtttaagtttaccgttaccgtacgtaggtaaggaaaggaaaggaacgaacgtaaacgtttaaccgtttaaaccgtaacgaaccgaacccgaaaccgtaaacggtaaaaacggaaaaaccgaaaaccgaaacgtaagtacgaacggaaacggtaacgtaacggaaggaaggaacgaaaccgtaaaacgttaaacgttaaacgtaaacgtaaaacgtaaacgtaaacgtaacctaaccgtaacgtaagttacgtacgtagtagtaacgtaacgaaacgaaccgtaaccgtaaacgtaaactaacgtaacgtaaccgttaccgttaccgttacccgttaacccttaaaccccttaacccttacgttacgtaacggtaacggtaaaggttaaagttaagtaacgtaacgtacgtacgtaaccgtaaccgttaaccgttaaccttaaccccgttaccccgttacccgttaccgttacggttacgttacgtaaccgtaaccgtaccgttaccgttaccgttacgtacgtacg
    >GRMETAB01AHC9E
    tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTAGGTGATGGAAACCTTTGGTAGACTCTTCTAAGTGCACGTGCAGATTTTCTCTTACCATGTATTTTACTTAAACAAATTTGTTTGTTATTTagtccacccgcctccttcttctacacacctctcgttccttcctaccgccactacttaacttagttgtcggttccccgttaccttcctaccgtccgttntgtcgtactanaccttcctgttgaccttgacgaagccacgaccgaagggactaggaggaggnngnnnnnnnnnnnnnnnnnnnnnnnnnnnnnnn

  • #2
    TTTNTTTNTTTTTTTTTNTTTTTTNTTTTTN looks like a poly A primer, broken up to avoid a huge homopolymer run.

    I'll also bet that the >GRMETAB01A4CCL read hit that wall of T's and died immediately, hence 'tatata'.

    Comment


    • #3
      Originally posted by GW_OK View Post
      TTTNTTTNTTTTTTTTTNTTTTTTNTTTTTN looks like a poly A primer, broken up to avoid a huge homopolymer run.

      I'll also bet that the >GRMETAB01A4CCL read hit that wall of T's and died immediately, hence 'tatata'.
      I still wondering whether the reads including poly-A primer are common or not. I mean which one is normal read?

      (1)
      tcagACGAGTGCGTAAGCAGTGGTATCAACGCAGAGTGGGAGAATCTCGAACCACATCTCTGCCCGTACCTAGCaacctaactctggtcgtnaagagctgntcncctcccncctcctntgtgtgtactactacgctcttntagacagcacgacgacacacggaggtaggagnn

      (2)
      tcagACGAGTGCGTGGGAGAATCTCGAACCACATCTCTGCCCGTACCTAGCaacctaactctggtcgtnaagagctgntcncctcccncctcctntgtgtgtactactacgctcttntagacagcacgacgacacacggaggtaggagnn

      Comment

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