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Genomic DNA extraction from filters


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#1 Pseudopneumo

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Posted 04 June 2013 - 01:21 AM

Dear all

I am wondering if anyone has experience in extracting prokaryotic genomic DNA (gDNA) from filters using a commercially available kit.

We have started trying to extract gDNA from air samples collected onto filters, using the PowerSoil kit from Mo Bio Laboratories, but have not had much luck; there was no sign of DNA as suggested on the NanoDrop.

Has any of you used modifications of this kit, or have used other kits to acquire gDNA? I have a feeling it is the low amount of microorganisms collected on the filters, but I am new to this project, and would like to just hear about different opinions regarding this issue.

Thank you very much.

#2 phage434

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Posted 04 June 2013 - 05:26 AM

It is very unlikely that you could see DNA extracted from air filters on a nanodrop. Typically, this DNA would be used in a PCR reaction with universal bacterial 16S primers, which were then purified, and either cloned for conventional sequencing, or end-tagged and used in high throughput sequencing.

#3 Pseudopneumo

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Posted 04 June 2013 - 02:26 PM

Thanks. We are going to use universal 16S primers for sequencing. So you are saying NanoDrop does not give an indication of whether there is DNA or not from these samples? I mean if there is no NanoDrop reading, chances are that there will be unsuccessful DNA amplification...?

Thanks

#4 phage434

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Posted 04 June 2013 - 03:09 PM

PCR can easily amplify DNA which is completely invisible on a gel or unmeasurable in a nanodrop. In principle, a single molecule can be amplified.




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