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  • eab
    Member
    • May 2011
    • 66

    #1

    Poor yield with chemical RNA fragmentation?

    Has anyone had problems with low RNA recovery after chemical fragmentation for mRNA seq? A collaborator says that he loses ~90% of his material when he fragments chemically. He says that covaris shearing of RNA provides a much greater yield. Has anyone else had the same experience?
  • upendra_35
    Senior Member
    • Apr 2010
    • 102

    #2
    Illumina kits uses chemical fragmentation for mRNA seq and they didn't see any problem with the yield of and so i don't think it is chemical fragmentation leads to low yield. Try to adjust the time of the fragmentation (typically 5 minutes).

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    • niceday
      Member
      • Apr 2010
      • 68

      #3
      What kit was used for chemical fragmentation?
      I used a commercial kit for fragmentation. The time was 15 mins. I tried fragmenting for 10 mins and increased it by 1 min up to 15 mins. I got the same result, all the RNA had been fragmented to below 100bp. I won't use this kit again.

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