Due to DNA extraction with phenol chloroform protocol, we have two different methods for using RNase as below:
image 1: 2% agarose gel without RNase
image 2: 1% agarose gel + 10 ul RNase
Is there any DNA smear in both of gels? or RNA?
P.S: target tissues were frozen kidney, liver and muscle.
Thank you in advance.
Mahmoud
I see - genomic DNA and probably degraded RNA.
If you will load to gell much less amount of RNA, you will see rRNA bands, but currently RNA degraded or overloaded.
thanks Ruslan,
Is there any possibility to make some disturbance in determination of DNA methylation rate, when degraded RNA exist?
Hi Mahmoud,
I agree with Ruslan. The bands at around 100bp should be degraded RNA. Assuming the loading amount is constant between with and without RNase treatment, you can see that the 100bp band intensity reduced after the treatment, indicating that it is likely to be fragmented RNA.
As for your second question, I think that it is heavily dependent on the technique you choose to analyse DNA methylation. Normally, bisulfite treatment followed by sequencing is commonly used. In this case, significant RNA contamination may contribute to notable portion of the sequencing reads. However, I have no experience in DNA methylation assay. Take it with a pinch of salt.
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