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  • mitochondrial sequences

    Hi Friends

    I know that mitochondrial sequences cannot be removed using any existing kits (Ribozero/Ribominus). How many sequences can be mitochondrial in any Illumina sequencing results? I sequenced few clones in my library using Sanger sequencing, most of them turn out to be mitochondrial and a couple of them rest but not rRNA. Do I need to optimize my cycle number? Any suggestion as to how should I try to optimize cycles?

    Deepak
    DPP

  • #2
    DNase treat your RNA samples prior to making your RNA-Seq library.

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    • #3
      thanks

      Originally posted by NextGenSeq View Post
      DNase treat your RNA samples prior to making your RNA-Seq library.
      Seems a nice suggestion but don't you think mitochondrial RNA will also interfere.
      DPP

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      • #4
        Probably, the worst library I've seen was 20% mtDNA (or mtRNA) and they didn't DNase treat their RNA.

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        • #5
          Thanks guys!

          Thanks for reply!
          DPP

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          • #6
            There is a nice paper in plos1 from Helicos that looks at rRNA and mitochondrial RNA content in different preps. polyA from brain reached 45% but most were around 10%

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            • #7
              Thanks

              Hey that is a good information. I did find the paper. Thank you once again.

              Bug

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              • #8
                Originally posted by The_Roads View Post
                There is a nice paper in plos1 from Helicos that looks at rRNA and mitochondrial RNA content in different preps. polyA from brain reached 45% but most were around 10%
                Link:Protocol Dependence of Sequencing-Based Gene Expression Measurements

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                • #9
                  That paper is 4 years old. Protocols have changed a lot since then.

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                  • #10
                    Originally posted by NextGenSeq View Post
                    That paper is 4 years old. Protocols have changed a lot since then.
                    I don't doubt that, but I am curious about how much mtRNA content you can expect in RNA-seq data.

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