The use of
Saccharomyces cerevisiae as a model system for molecular biological research has always been popular. The ease of laboratory culture and manipulation, including the ability to perform transcriptional and translational studies, makes yeast an ideal system for research. Unfortunately, the current protocols for DNA extraction in yeast are both time-consuming and difficult. The reason is that yeast cell walls are hard to lyse – existing protocols to do so requires the use of harsh reagents and rough mechanical lysis, both of which are potentially damaging to the DNA. The Y-DER Yeast DNA Extraction Reagents from Pierce allows for DNA isolation without dangerous reagents or glass beads and little to no RNA contamination, all in about one hour.
The Y-DER Reagent is optimized for use on Saccharomyces cerevisiae, but can be used with other organisms. DNA can be obtained from single colonies, fresh or frozen cells or cell pellets from liquid cultures. The proportions in the protocol must remain constant, but can be scaled up or down depending on the researcher’s needs. I have used the Y-DER Reagent to purify large-scale Saccharomyces cerevisiae cultures (1 L) separated into approximately 700 mg cells/tube. I scaled the protocol up quite a bit and collected the eluted DNA at the final step.
The Y-DER Reagent is a tremendous benefit to those interested in purifying yeast genomic or plasmid DNA. Anyone who has used a beads beater or lysed cells by hand with a vortex will appreciate this kit. Also, omitting the use of chloroform and phenol is a plus. Obtaining DNA from a single colony or from a liquid culture widens the applicability of this protocol. Even though genomic DNA is purified with plasmid DNA, the sample can be directly used for PCR and the protocol gives information on how to optimize use for bacterial transformations. I have used the purified samples for transforming electrocompetent bacterial cells and have had success (i.e. a high number of transformants).
The only drawback that I have run into with this protocol is that after an EtOH precipitation step, there can be a large amount of what appear to be salt remaining. To deal with this, I generally wash the pellet multiple times with 75% EtOH, instead of the one time that the protocol suggests. Because this could mean going through lots of reagents, Pierce does offer the kit in bulk size, which is 50 times larger than the regular kit, but of course this is more expensive.
Overall, the Y-DER Yeast DNA Extraction Reagent is an improvement to current methods of yeast DNA purification. It enables the researcher to purify DNA quickly and easily, allows for scaling the protocol to your specific research and produces a product that can be used for multiple tasks down the line. The kit is relatively inexpensive and can be purchased in bulk.
Karrie Comatas
Research Technician II
Duke University