Last week I submitted 6 organisms to the UCD Sequencing Facility after several weeks of isolating and purifying their 16S genes. My initial submission of 5 organisms went somewhat smoothly, though something managed to sneak into two of my samples and mix the forward sequences I received (for each organism, we must submit enough for both a forward and a reverse reaction to be sequenced. We receive these sequences and align the complementary bases to generate a consensus sequence). I had really hoped this time would go much more smoothly than last, but unfortunately 4 of my 12 reactions, all forward reactions, were mixed and thus indecipherable.
I suspected the forward primer might have been contaminated at some point, and looking at my initial submission the only mixed reactions were forward reactions as well. Seems pretty clear to me that some little critter was able to find his way into our forward primer and rain on my alignment parade. I had hoped that I would have a larger list of organisms by this point, but here is what I have analyzed so far
- Staphylococcus pasteuri
- Enterobacter ?
- Bacillus amylolequefaciens
- Micrococcus luteus? (the most “interesting” organism I’ve found, based on GOLD results)
I brought in 5 new samples that I can hopefully begin culturing in the coming days, and in the near future I will resubmit the forward reactions that failed using a new batch of forward primer.