I have extracted RNA from fungus using Trizol method. What are the extra bands in gel image(1) other than 28srRNA AND 18srRNA? The 260/280 ratio of samples were 2.05 and 2.14 range .Can i proceed with these samples for transcriptomic studies?
Several times I have extracted RNA from the same fungus using the same protocol. If here RNA quality is fine there was gDNA contamination (image 2). If there was no gDNA contamination RNA quality was not good. I have treated samples with Dnase I as recommended by manufacturer ( 2.5microlitre Dnase upto100microgram of RNA sample, Rnase free Dnase set from qiagen). But DNA not get degraded.
Can I add Dnase in between RNA extraction as in DNA extraction after aqueous phase transfer?
Can anyone help me?
Hi,
The extra band is gDNA contamination. How long you are treating your RNA with DNAse I?
For checking the integrity of your RNA you have to use formaldehyde gel and run in 0.5 x MOPS buffer.
From what you show here ( actually the total length of the gel would have been more informative to see the small RNA species and degradation as well as nucleic acids stuck on the top if any). You must check you DNase activity on a plasmid. You may also have RNA degradation.
- Badges
- Science topic
- Similar topics
- Analytical Chemistry
- Extraction