I have been using Qiagen's DNeasy kit for DNA extractions of liver flukes preserved in 100% ethanol. Once the lysis buffer and proteinase K are added the solution gets a strongly tanned brown color even with the tinniest piece of tissue. It stains the column mesh brown also and after the extraction protocol is completed, I end up with little or no DNA with very poor quality according to the nanodrop. Is there anyway to prep these samples prior to a kit extraction protocol?
Hello, try 70% ethanol instead of 100 keep one more day and than start extraction.
You can also try to add 1% PVP (PolyVinyl Pyrrolidone) either while grinding the sample or in the lysis buffer to get rid of polyphenols and polysaccharides. I have successfully extracted high quality DNA from some marine species like polychaetes, clams, and fishes Andrew Davinack
Hi all, thanks for the info. I've successfully amplified high quality DNA from several species including polychaetes, fishes, ticks, molluscs, arthropods, etc. But for some reason the adult trematodes are giving a lot of trouble. I will try using those methods you suggested and see what comes of it.
Thanks!
Hi Andrew !
I have been working for some time with similar flukes (Fasciola hepatica) and it works just fine for samples stocked in 80% EtOH. Also, we do the DNA extraction with Chelex and for PCR we usually have to dilute the samples because there is too much DNA. In my opinion, you have to make sure you have retired all possible ethanol from your sample prior extraction. Hope this can help! Cheers, Antonio
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