Hey folks,
I am trying to design an experiment to obtain DNA from historical specimens of frogs (some of them were collected >100 years old.
Anyone experience with the manufactured DNA extraction kit from FFPE tissue samples? Could you share your experience in using this kit compare to the normal tissue extraction kits or the old-school methods (e.g., phenol-chloroform method)? Which one work best (e.g., produce large amount and high quality of DNA required for subsequent steps)?
Also, what do you think about this study?
Article DNA extraction from FFPE tissue samples – a comparison of th...
And if anyone experience with the forensic extraction kit?
I really appreciate your help!
Best regards,
Hi Umi, for historic samples we use phenol-cloroform extractions but use Amicon Ultra 0.5 mL filters during the purification step instead of ethanol precipitation. So far it worked well, we've extracted DNA from pretty old specimens (more than 150 years old). However, we don't know if the dry skins or wet specimens we extracted the DNA from had been fixed in formalin. Good luck!
I have experience with extracting DNA from FFPE tissues but the problem was that I obtained degraded DNA with high quality , then we found that the degradation was because of the concentration of Formol . I tried 3 protocols (phenol, salting out and Kit quagen).
Find attached Salting out protole and a book will help you .
Good luck
Thank you Arlo Hinckley and Hossam Hilal el Idrissi !
I agree that it is very possible to extract DNA from FFPE, and it makes your life much easier if you know the fixation and preservation history of the specimens. Formalin may result in DNA damage such as methylation and cross-linking, but it probably also protects the DNA from hydrolysis (in which the DNA basically just breaks down into individual nucleotides).
Hey Hossam, would you mind sharing your experience using the 3 protocols you mentioned? Did you use the same size of tissue for the three protocols? And how was the results? Which one yield more amount of DNA? Which one produce higher quality of DNA?
Hi Umilaela Arifin .
Three methods of DNA extraction from 18 FFPE samples (for each method) were tested: salting-out; Phenol-chlorophorm methods and EasyPure® commercial kit. Two common methods have been used to measure quality and quantity, DNA purity / concentration is measured for quantity using a spectrophotometer (absorbance) and quantity using an agarose gel analysis.
The best concentrations and purity of DNA were obtained by salting-out method and Phenol-chlorophorm . The BCL-2 gene was amplified by PCR to evaluate the equality of extracted DNA. There was no amplification for salting-out and phenol chlorophorm methods. Instead, the BCL-2 gene was amplified only in 2 of 18 samples extracted by EasyPure ® FFPE kit.
Size: We used the same size of tissue for the three protocols, 10µm.
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