mitochondrial sequences

[deepakpatilp]

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

[NextGenSeq]

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

[deepakpatilp]

thanks

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.

[NextGenSeq]

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

[deepakpatilp]

Thanks guys!

Thanks for reply!

[The_Roads]

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%

[biochembug]

Thanks

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

Bug

[BrianU]

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

[NextGenSeq]

Link:Protocol Dependence of Sequencing-Based Gene Expression Measurements

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

[BrianU]

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.