1
$\begingroup$

I am going into the field soon to sample a very remote environment. For reasons that are unimportant here, I would like to extract bacterial DNA from these samples in the field.

I will have access to electricity, but no heavy equipment such as centrifuge. My goal is to extract DNA from these hypersaline liquids for long-read sequencing and subsequent metagenomic analysis with a focus on DNA methylation. My plan is to filter large volumes to have sufficient biomass onto a filter from which extraction will occur (think seawater type samples).

Any suggestions for an equipment-less DNA extraction kit? Vortexing is fine.

$\endgroup$

1 Answer 1

2
$\begingroup$

You'd be far far better off waiting until you can get them to a lab. Given the sensitivity of the technique and the unknown abundances and likelihood of getting samples, giving them the best extraction you can is the way to go. I realize this isn't always feasible and there are reasons not to do an extraction in the lab, but it is generally better. Fixing in ethanol on a filter would be fine to preserve until you can get it to a lab.

It would depend on the scale of preparation you want to do, exactly what you want to get out of it and the abundance of the bacteria in your samples. There are a number of para-magnetic bead based DNA (AKA SPRI beads) and RNA extraction products on the market. These are highly sensitive and quite quick to perform, but you do need the magnets and beads, both of which are quite expensive.

An example of this used for environmental bacteria is the following article. They did their extraction in the lab and used a centrifuge to concentrate samples, but it could easily be applied to the field:

Byrne RL, Cocker D, Alyayyoussi G, Mphasa M, Charles M, Mandula T, Williams CT, Rigby J, Hearn J, Feasey N, Adams ER, Edwards T. A novel, magnetic bead-based extraction method for the isolation of antimicrobial resistance genes with a case study in river water in Malawi. J Appl Microbiol. 2022 Nov;133(5):3191-3200. doi: 10.1111/jam.15755. Epub 2022 Aug 23. PMID: 35946113; PMCID: PMC9804433.

$\endgroup$
2
  • $\begingroup$ The reason to do it in the field is to avoid changes in the methylation pattern changing due to temperature shock when storing. One thing I may do is flash-freeze after storing in a buffer instead, but I did hope for a baseline to compare. Is ethanol the recommended storage buffer? $\endgroup$
    – donkey
    Mar 27, 2023 at 4:07
  • $\begingroup$ @donkey ethanol more or less fixes the DNA as it was at the time of addition. I don't know off the top of my head how you would proceed to with an extraction once in ethanol, but it should be fairly easy to do. EtOH fixed samples are one of the better sources for DNA from historical museum specimens and the like. $\endgroup$
    – bob1
    Mar 27, 2023 at 7:55

You must log in to answer this question.

Not the answer you're looking for? Browse other questions tagged .