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dna extraction from soil samples


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#1 alkb

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Posted 11 May 2010 - 12:32 AM

Hello all!

I am new in this forum. Sending sunny greetings from Germany... :P

I started to work in terrestrial ecology field few weeks ago. My project is to asses microbial diversity in polluted soil using qpcr. But I already faced first problem, which is DNA extraction. I tried with protocol they usually use in the lab for "normal" soil. The concentrations were very law, the most problematic is 260/230 ratio (around 0,7 -1,4). I also tried with NucleoSpin Soil Kit, results pretty the same...

I need ideas, advices... how to isolate relatively pure DNA with reasonable concentrations from soil, enivronmental samples

Thank you in advance.

#2 gebirgsziege

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Posted 11 May 2010 - 01:02 AM

have you tried a pcr with your DNA and checked the yields/quality of the result?

Maybe the bad reading comes from something in the polluted soil interfering with the photometric reads. I have the experience that with soil these photometric and quantifications methods are only partly reliable.....too many factors that can influence your results...
A man cannot be too careful in the choice of his enemies. (Oscar Wilde)

#3 alkb

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Posted 11 May 2010 - 02:01 AM

have you tried a pcr with your DNA and checked the yields/quality of the result?

Maybe the bad reading comes from something in the polluted soil interfering with the photometric reads. I have the experience that with soil these photometric and quantifications methods are only partly reliable.....too many factors that can influence your results...


Thanks for your answer. No, I didnít run pcr yet, because I though good dank quality is a "must be"!!! :P

#4 gebirgsziege

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Posted 11 May 2010 - 02:37 AM

It is good if you have, but I had horrible samples which were working perfectly, so I would give it a try.
A man cannot be too careful in the choice of his enemies. (Oscar Wilde)

#5 Maddie

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Posted 12 May 2010 - 05:39 AM

I think humic acids have the same absorbance as DNA, so your concentrations might be totally wrong.
You can always add a cleanup step if you want with a Qiaquick or MinElute.
Try the PCR and don't forget the BSA.
Theory is when we know everything and nothing is working. Practice is when everything is working and nobody knows why. Here, we combine theory and practice. Nothing is working and nobody knows why.

A. Einstein




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