Summer Session-Early PCR Success

Hello! Welcome back.

Last week, I got promising results after testing two simple thermal DNA extraction methods on my gram positive species. The gels I ran post-extraction were positive for DNA banding.

Protocol #1 utilized an SDS/TBS mixture and boiling to lyse the cells and took approximately 45 minutes to set up. Protocol #2 used sterile water and boiling and took about 20 minutes from start to finish.

Bolstered by the results from both tests, I took the DNA samples obtained using protocol #2 and decided to run a quick PCR on them using a primer set that I designed last semester. As you may recall, I was able to amplify DNA from 8 out of 9 targeted gram negative species using a primer set of my own design; this experiment capped off my work last semester.

I fully did not expect to get positive results from this PCR, as my primers were designed using three gene sequence maps specific to three of the gram negatives I previously used for this study. However, after repeating the reaction using six gram positive species, DNA was successfully amplified. The gels I ran post-PCR are shown below.

This is great news! I can now move forward with the next phase of this study. Details on that to follow in a later blog.

Until then, live long and prosper.

Top: Gel 1 Bottom: Gel 2. Both gels were run using the same DNA samples. There are 6 wells loaded; from left to right the species are B. subtilis, S. aureus, B. cereus,S. epidermidis, E. faecalis, and M. luteus. Gel 1 used 2 µl of loading dye and 2 µl of sample; gel 2 used 2 µl of loading dye and 4 µl of sample.