When you’re working with radioactivity it seems like the simpler the labeling system the better. Fumbling around trying to open multiple micro-centrifuge tubes when you’re double-gloved, making sure that you’re not dripping label all over the bench is just not a pleasant experience. The people at GE Healthcare must have realized this and their solution to this problem is the Rediprime™ II DNA Labelling System. This system contains everything that you need to generate a random-primed labeled probe in one tube - all you need to do is add the template and the label and you’re ready to go.
The Rediprime II system is set up to generate dCTP labeled probes (using either 32P or 33P labeled dCTP) in less than 20 minutes. The reagents for generating a random-primed probe are pre-aliquoted and freeze dried (for stability) in 1.5ml tubes (including the Klenow). This kit can be stored at room temperature and when you’re ready to label your probe, you simply denature your template (boil and quick cool on ice) and add to the tube along with your label (of course GE Healthcare recommends using their Redivue™ label). The reaction can proceed for as little as 10 minutes at 37° C or, if you’d like, you can let it proceed at room temperature for 20-30 minutes. Because there’s no need to remove unincorporated label in a random primed probe, once your labeling time is up, you’re ready to hybridize.
I have used this system to label a countless number of probes and have never had a problem with it. One worry is that the Klenow could go off after being stored at room temperature for a long period, but I have used kits that are more than 4 months old and haven’t had a problem with labeling. In the product specs GE Healthcare claims that this system can be used to label as little as 25 ng of DNA and in my experience this is true. Although it’s not necessary, I have purified the probe away from the unincorporated label and have been pleased with the specific activity of my probe. They also claim that you can label DNA in low-melt agarose, although I have not tried this. The only downside to this product is that since the unlabeled nucleotides are already in the tube you have to use labeled dCTP (which, to be honest, isn’t much of a downside).
All in all, I would have no problem recommending the Rediprime II™ DNA Labelling System. It’s fast, easy to use and has worked for me everytime I’ve used it. No one enjoys working with radioactivity and this system can cut down on the amount of time that you have to spend at the hot bench.
Peter Haggie, Ph.D.
Post-Doctoral Fellow
University of California, San Francisco