Tuesday, January 13, 2009
Here in the Bay we purify a heck of a lot of plasmid DNA using over-priced commercial kits. Mainly because there's a fully stocked Qiagen biobar down the hall and this set-up appeals to our excessive laziness. But sometimes even laziness can have its limits. Maybe the Ottawa transit strikes and blizzards are getting to me. Whatever the reason, today I was particularly struck by the ridiculousness of shelling out hundreds of dollars of grant money for a week's supply of aqueous solutions and mass-produced plastic tubes, packaged in totally unnecessary cardboard and throwaway Nalgene bottles. Surely a Ph.D. biochemist can come up with something more efficient than this?
The simple two-step procedure used by Qiagen and most other commercial plasmid prep kits is no secret. First comes Birnboim's alkaline lysis - a series of three buffers that lyse the bacteria and then neutralize the lysis reaction. Second, plasmid DNA is purified from the lysate by passing it over a silica column in the presence of high salt. Making the buffers is trivial (see below). Plus, we probably have a lifetime's supply of unused buffers from previous Qiagen kits lying around the lab. So we're paying for the little plastic columns. About $1.60 CAD a pop. Making the columns is probably not much fun, and alternative materials don't seem to be as efficient. But in theory there's no reason the columns can't be re-used.
The idea of regenerating commercial miniprep columns first came to my attention a while back. This paper/ad described a two-buffer product called MaxxBond that allowed columns to be regenerated and re-used up to ten times. I liked the idea of recycling columns, but not the idea of relying on yet another over-priced, proprietary/patented technology of an unknown nature. I was particularly concerned about the possibility of unknowingly cross-contaminating my plasmid preps, which would be a total nightmare. It's impossible to assess the risk of this happening without knowing how the columns were being regenerated.
After a little discussion in the Bay today, a quick surf on the Google produced a better alternative now floating around the interweb. The Indian research group of Ranga describe a method that involves simply storing the columns in hydrochloric acid and rinsing them with water when you're ready to use them again. They claim commercial columns can be re-used 10-20 times in this way. What I find appealing about this method is that the HCl acid works by physically destroying residual nucleic acid from your last prep, so it would seem pretty much impossible to cross-contaminate the next one. And a 10-fold increase in column productivity sounds pretty darn nice, and drops that price from $1.60 to 16 cents per prep. I think I might actually try it.