I would like to know which DNA extraction kit would work best for extracting bacterial DNA from animal tissue, specifically the pleopods of a crustacean. Some people have mentioned the DNeasy blood and tissue kit has worked for them, while others have suggested the power soil kit. I understand a kit with beads to aid in bacterial cell lysis is preferable, but any advice would be appreciated.
The best kit for this application is the
Thermo Scientific™ Genomic DNA Purification Kit Cat# FERK0512 (product instruction sheet attached). Follow the instructions for "tissue" and freeze the pleopod in liquid nirogen, grind to a fine powder, and follow the instructions provided. The 65 C incubation in step 1 of the protocol may be extended to up to 30 minutes with agitation to increase extraction of DNA. The lysis solution will do the job of lysing the bacteria - no need for beads.
Good Luck!
hi
The Qiagen company products are highly efficient in the field of DNA and plasmid extraction.
"The DNeasy Blood & Tissue Kit from QIAGEN"
http://www.scielo.br/pdf/cr/v46n7/1678-4596-cr-46-07-01223.pdf
good luck
I think if you use Liquid Nitrogen or sonication, you can easily use high quality kits but not necessarily soil kits that are very expensive.
Even you can use manual methods.
Regard.
With all respect to my colleagues Reza Hadavi and Alireza Mordadi The DNeasy Quiagen Kit is not the best for this application. The DNeasy kit relies on PBS and proteinase K to lyse cells and this is not efficient for bacteria nor is it sufficiently harsh to recover DNA from bacterial cells that are adherent to the pleopod surface. The Kit I mentioned above uses a strong denaturing solution to lyse the sample and will be superior for extraction of DNA from bacterial cells and hard tissues such as crustacean exoskeleton. In fact, the paper cited by Reza Hadavi shows that a genomic DNA isolation kit similar to the one I mentioned above performed as well as the Qiagen kit. While the Qiagen kit is an excellent product and I have used it myself, as someone who has experience in extracting DNA from arthropod samples specifically I can tell you that it will not be as effective in this application. The paper cited is extracting DNA from bovine soft tissue, and that is a much different matrix than crustacean pleopods. April Stabbins original question was what method is best for extraction of DNA from crustacean pleopod tissue. Simply stating that a variety of techniques exist or recommending a kit or method that hasn't been validated for the application in question is not helpful. Please kindly answer questions about which you have genuine expertise and in a manner that addresses the issue of the original poster.
Regarding to your answer, @Zachery R Belak.
Thanks for a very clear explanation. I am in the same page with you as I mentioned first using a harsh methods like liquid Nitrogen for breaking cells and then using such kits.
Please accept my apologies for unclear explanation.
Regards
Thank you all for the information, that's really helpful! I think what Zachery R Belak has suggested seems the most appropriate and given you have had previous success using those methods with arthropods I will follow that protocol first. Thanks again!
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