Although I’m not involved in the marketing side of research kits and the like, I would imagine that getting scientists to put down their tried and true methods for even a short time to try something new must be very difficult. This task is especially challenging for those areas, such as plasmid isolation, that have been dominated by one company for many years. As difficult as this may be, it’s always a good strategy to approach the potential user from the universally understood angle of money, i.e. offer a free trial kit. This is exactly what Promega (Madison, WI, USA) has done with its new PureYield Plasmid Midiprep System. You can go to their website at www.promega.com and request a trial kit of 4 preps gratis.
I used the PureYield kit for my last plasmid isolation and was very pleased with the whole experience. In addition to being very simple to use, all reagents can be stored at room temperature. So, no more looking through the fridge for the one buffer that must be stored at 4°C, and then wondering how old it is. All the PureYield reagents are right there in the box. Four of the six solutions are supplied ready to use; one requires the addition of ethanol and one isopropanol. The general scheme of the kit is similar to most other kit-based methods: lyse the bacterial pellet, neutralize, clear the lysate, bind the DNA to a matrix, wash, and then elute the purified plasmid. An added benefit to the Promega kit is the option of using only centrifugation for the various steps, only vacuum filtration, or a combination of centrifugation and vacuum filtration. All methods are equally straightforward but the vacuum-based method probably cuts 25 min or so off the time of the procedure. The all-vacuum method combines the lysate clearing step with the DNA binding step by cleverly allowing the clearing column to be nested inside the DNA binding column. The lysate is cleared and the DNA bound to the matrix in less than 10 sec.
Another plus of the kit is its wide range of suggested bacterial culture volumes. A standard protocol is provided for cultures of ≤ 50 ml and an alternative method that employs a centrifugation step to preclear the lysate prior to column clearing is provided for cultures of 51-250 ml. The ability to purify plasmid DNA from cultures as large as 250 ml results from the 400 µg DNA binding capacity of the matrix.
Of course, none of these benefits would mean anything if the kit didn’t work well. Fortunately, at least for my initial trial, it worked beautifully. From a 50 ml overnight culture of DH5alpha, I recovered 50 µg of DNA with an A260/A280 ratio of 1.9. Not bad at all.
So, how much does all this convenience cost? Not much at all actually. The PureYield kits range from $6.55 to $7.20/prep, depending on the number of preps in the kit. In contrast, comparable kits from the leading brand cost around $18.00 to $24.00/prep, depending on the number of preps and whether you want the ‘fast’ variety, or ‘endotoxin free’. By the way, the PureYield kit includes an endotoxin removal step, and is compatible with both EndA- and EndA+ strains.
Michael Campa, Ph.D.
Asst. Research Professor of Radiology
Duke University Medical Center