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Ten grams of DNA

Scott Chimileski-DNA extraction- Haloferax volcanii

Ten+ grams of crude DNA, extracted from 5 liters of Haloferax volcanii culture.

Almost all biologists extract DNA for one reason or another. But for just about every routine application, plasmid preps, sequencing, and the like–we are used to dealing with nanogram or microgram concentrations–delicately resuspended from the most minuscule pellets, barely visible on the side of a microtube. This is not a tale of nanograms or micrograms.

For my first project as a PhD student, I studied the idea of extracellular DNA (or eDNA) as a nutrient in haloarchaea: a group of “extremophilic” archaea that live at very high salt concentrations. We knew from population genetic studies that haloarchaeal species were promiscuous in natural hypersaline sites, able to readily exchange genes horizontally (that is, independent of vertical inheritance through reproduction). But it was unclear how they were doing this: which actual cellular mechanisms mediated gene transfer. One possibility was natural uptake of eDNA, a process that had not been studied in haloarchaea (or essentially any species within the domain Archaea for that matter). So, inspired by discussions with my advisor and by earlier work on eDNA as a nutrient in bacterial species, I began my investigation of DNA uptake by asking whether the species Haloferax volcanii could “eat DNA.”

I needed DNA to provide as a possible growth substrate–a lot of DNA. I quickly found that typical DNA extractions were not going to be of much help. I was testing many replicates, DNA isolated from a range of species, several related conditions, and trying to figure out whether DNA could act as a carbon, nitrogen and/or phosphorus source (after all, DNA offers all three of these essential elements of life).

I did what any scientist would do: I scaled-up … and scaled-up, and scaled-up. The fact that I needed highly purified DNA meant I needed even more to start with, because of course some is lost with each purification step. I finally ended up with a purification process that began with an ethanol extraction from 5 liters of spun-down liquid culture. Adding the ice cold ethanol in 2 liter flasks was always exciting at the end, when gobs of DNA (a bit of RNA too) would precipitate instantaneously: a slurry that you could probably spool with a baseball bat. Even after several subsequent phenol-chloroform extractions to remove contaminating protein, RNA digestions, and removal of low molecular weight fragments and other impurities, I was able to yield several grams of pure DNA.

I share these photographs from one of my extractions back in 2010, as I wonder whether many others have seen so much DNA in one place. Beyond literally intending to use it as “food,” it is difficult to imagine other experimental contexts requiring grams of DNA.

This work ended up leading to into a great story that is still unfolding. Extracellular DNA turned out to be a nice thread (no pun intended) into my next project too: considering it is also component of haloarchaeal biofilms.

 

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