Like a few other people in the lab, I’ve been struggling with DNA extractions of late. I’ve tried several methods, including the “columnless” method that a few people are using. But I was having pretty spotty luck. Then, during a bout of methodological soul-searching, I also tried a CTAB protocol that used to be my go-to method. It failed me a few weeks ago, but in a second trial last week, it delivered the goods: loads of very pure, high molecular weight DNA. It might not work for all plants, and there do appear to be situations in which it is not the best option, but if anyone wants to get back to down-home, from-scratch extraction, the way momma used to do it, then give this one a try:
Please note: the results below are for Streptanthus (the plant I’m studying; Brassicaceae); I’m trying it for some Asteraceae right now, and will update the post once I have results.
Momma’s DNA Isolation via CTAB
It’s pretty basic, and I think that might be the advantage that it has over other, fancier methods. Overall, I’ve found that the single most important factor in the success of this method is the amount of tissue used. Less is definitely more, as they say. For plants that are fleshy or succulent, use less tissue. If using dry tissue, never use more than 20 mg. I have recovered as much as 40 µg of DNA from 20 mg of dry tissue, so you can err on the side of less tissue and still get plenty of DNA.
Here are some results that I obtained using 60 mg of fresh leaf tissue. This is with a 100 µL elution, so I could have as much as 30 micrograms of DNA from this extraction. The Nanodrop profile is for one of two samples; the other had about half this amount of DNA.
Here is the gel; the sample above is the fourth one from the left.
If you want to try it, let me know and I can lend reagents or run a few of yours myself.