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  • RiboZero won't work?

    Hi,

    I'm trying to make a library of RNA following the ScriptSeq protocol. Everything works just fine except the RiboZero depletion of rRNA. I'm using young leafs of rice. An input of 2.5 µg of sample, 8 µl of removal solution.
    I have to do this step twice and even then, I don't get rid of all the rRNA. I still have some nice peaks on the bioanalyzer.

  • #2
    I know many kits have very specific maximums and just increasing the volumes doesn't always work.

    I know Epicentre has a max of 1ug and over that causes incomplete reduction.

    Not familiar with Script Seq but I would suggest checking the maximum amount and doing multiple replicates if you are over the maximum.

    Comment


    • #3
      Originally posted by kwaraska View Post
      I know many kits have very specific maximums and just increasing the volumes doesn't always work.

      I know Epicentre has a max of 1ug and over that causes incomplete reduction.

      Not familiar with Script Seq but I would suggest checking the maximum amount and doing multiple replicates if you are over the maximum.

      According to their protocol the input should be between 1 and 5 µg. I'm using standard 2.5 µg so that cannot be an issue

      Comment


      • #4
        Hi Neiltje, Which Ribo-Zero kit are you using, and what is the sample type (human/bacteria/yeast/plant)? RZ can use up to 5 µg input total RNA, and depending upon the Ribo-Zero kit used, as little as 100 ng.

        Comment


        • #5
          Originally posted by Olaf Blue View Post
          Hi Neiltje, Which Ribo-Zero kit are you using, and what is the sample type (human/bacteria/yeast/plant)? RZ can use up to 5 µg input total RNA, and depending upon the Ribo-Zero kit used, as little as 100 ng.
          I'm using the RiboZero plant leaf on young leafs of rice. I think it just doesn't work well on this material because the last time I tried, I also took a sample of Canola and that worked fine

          Comment


          • #6
            Variations in rRNA sequences even between some species can change the outcome. Do you have the rRNA sequences for your strain of rice?

            Comment

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