I did DNA extraction using Qiagen DNeasy Plant Mini Kit. I ran out of RNaseA and I did not add RNaseA in couple of samples.
Nanodrop reading is very different for samples with and without RNaseA.
Other samples have concentration ranging from 12-45 ng/ul
A260/280 ranging from 1.84 - 1.97
A260/230 ranging from -3.47 to 10.30
For samples without RNaseA, Concentration 213 and 275 ng/ul
A260/280 2.1 and 2.22
A260/230 2.55 and 2.47
Why is concentration so high on samples without RNaseA?
Do these A/260/280 and A260/230 values looks good?
Can these samples be sent out for genotyping? will there be any issues with genotyping?
RNA comprises the majority of the nucleic acids in a cell, which is why the samples without RNase treatment show a much higher concentration.
In theory, your genotyping reactions could still work since they will be using a DNA-dependent DNA polymerase, but you won't be able to accurately determine how much DNA you are loading into your reactions.
RNase uses to remove your RNA from your samples. DNA extraction with RNases will result in removing RNA contamination.
You can buy RNases separately from Qiagen.
Your samples should be just fine for genotyping. Yes, having the RNA present will make it challenging to get an exact quantity of nucleic acids but polymerases are template specific. Also, the amount of gDNA that can be used for PCR has a wide range of working concentrations.
(I'm assuming this is a standard PCR-based genotyping)
The only issue I've had with PCR on gDNA samples is from species with lots of secondary compounds present, such as sweetgum tree leaves (lots of resin)
Good luck!
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Biomolecules
- Nucleic Acids
- RNA