I tried DNA extraction from brown algae, i used to digest the tissues by using lysis buffer (0.25 mTris borate, 0.1 mEDTA, 2% SDS, and 0.1 mNaCl set at pH 8.2) instead of using liquid nitrogen but I unable to get the DNA by the method which I followed. Any one can suggest alternative method for DNA extraction.
Hi,
Please, see the attached document in which the researcher used a protocol of (GENOMIC DNA ISOLATION FROM GREEN AND BROWN ALGAE), which may help you to get a required yield of DNA with good quality.
Good Luck!!!!
Dear Dr.Eminem thank you for your help and initially i have tried to followed this technique but i don't have the 50–55 mm nylon mesh so that i have tried another method. Any way i will try to get the mesh and again go for this technique .......
Try this method "A fast and inexpensive DNA extraction/purification protocol for brown macroalgae" (open article) http://onlinelibrary.wiley.com/doi/10.1111/j.1471-8286.2006.01587.x/abstract
Good luck.
try
http://eprints.lib.hokudai.ac.jp/dspace/bitstream/2115/52032/1/JAP25-1_337-347.pdf
I have used a CTAB procedure for over a thousand extractions from mostly fucalean brown algae for PCR. My procedure is much the same as in the Varela et al 2006 paper suggested by Hemin but without the preliminary nuclei extraction step.
I just grind up a silica gel dried sample in CTAB with a micropestle then proceed to the incubation and purification steps. I use 1% PVP. I have added b-mercaptoethanol but haven't noticed any improvement. Usually I use two CIA purification steps (adding Chloroform/isoamyl, shaking to emulsify, centrifuging then removing the supernatant, then repeating).
My experience is that sample selection is important - use a very small quantity of clean tissue - I use the top 2mm (at the most) of the (dried) growing tip of fucalean algae. This should mean more DNA (small young cells) and less secondary compounds.
I have used this method successfully in Sargassum, Cystophora and Carpophyllum as well as other brown algae.
I have used liquid nitrogen when I want a large amount of DNA for particular applications, but grinding in CTAB works fine for PCR.
Dear Joe Buchanan,
Exactly correct his methodology works but the amount of DNA i got is very less and i hope some protein may not wash properly during the last two wash steps. I will go for same method with slight modification and will see how it works.
Thank you guys for your valuable information now my methodology works very well...............
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