I am using Zymo Quik DNA FFPE Tissue Kit for DNA extraction; I followed the manufacturer's protocol, but I've seen a smear. What should I do to avoid it? Should I proceed with PCR amplification? I have attached the picture of DNA from ffpe tissue after electrophoresis .is it what I suppose to get?
you can proceed with PCR if your purity ratio was between 1.8 to 2 and do that for few samples first and check the PCR product with GE
however, I prefer either you repeat the extraction or change the protocol
for FFPE I always prefer the conventional way like what they do here
Article DNA Extraction from Paraffin Embedded Material for Genetic a...
see the video
Good luck
Hello,
After DNA extraction it is essential to run the DNA on an agarose gel to check the quality of DNA. A tight band of high molecular weight indicates a good DNA, while smearing will indicate degraded DNA.
DNA of high quality is needed for further analysis. I would suggest you do not go ahead with PCR but rather repeat DNA extraction.
Good Luck.
It appears that the DNA is degraded, so I would not run PCR on those samples.
Historically, extracting DNA from formalin-fixed tissues has been problematic. Formalin exposed to air readily oxidizes to formic acid, which will cause depurination of the DNA. So if the formalin used to fix the tissue was not fresh, it probably degraded the DNA.
I would get fresh samples and make sure the formalin used to fix them is fresh.
- Badges
- Science topic
- Similar topics
- Biological Science
- Anatomy
- Tissue