Hi, I am extracting total RNA from plant roots. I am getting low concentration of RNA (~250ng/ul) and the ratio of 260/280 is 1.74. I already have isolated RNA from the same sample previously and got good result. But can't understand what's wrong this time. I am using Trizol and Qiagen RNeasy plant mini kit for RNA isolation. Can some one suggest something?
The concentration can vary because two factors mainly: amount of tissue or cells that you are extracting RNA, and dilution (volume of diluent where you diluted the sample). Did you run a integrity gel of your sample to see if the RNA is degraded?
RNA extraction main problem is during purification. If you use a spin column, in the last step do ilution for 2 or 3 times.
The obtained rna should be diluted in low amount of rnaase free water
Yes we repeat the elution procedure twice and we elute in 30ul RNase free water.
Thank you everyone for replying. We store samples in -80 freezer. Similar procedure is followed for each samples. We use Qiagen kit for RNA extraction. The buffers are working fine as we do not face any problem while extracting RNAs from other samples. This is happening with particularly this root sample. Is it possible that the phenolics present in the samples are somehow inhibiting the procedure??
Thank you very much for this, We assume that phenolic compounds can be the reason for getting such result. And also the root sample is also old.
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