The biotechnology industry is one of the most dynamic out there – in fact, it never stands still! For non-scientists this can be intimidating. For scientists, it can be challenging to explain what we do in ways that non-scientists can understand and appreciate.
Scientists have made great strides in improving our ability to use molecular processes to our advantage, from discovering the basics of how to isolate and manipulate DNA to gaining an understanding of how genes direct the creation of proteins in cells. It’s clear that there is a lot we can contribute to the scientific literacy of the general public.
Since the seminal paper on bisulfite conversions is celebrating its 20th anniversary this year, I thought it would be nice to feature, over the next few weeks, how the bisulfite conversion technique came to be and the assays that have been developed around this seminal technique.
How It All Began…..
In 1970, Hayatsu et al. discovered a chemical interaction between sodium bisulfite and pyrimidines that would have a tremendous impact on how DNA methylation patterns and changes are studied. This group found that uracil, thymididine, and deoxycydidine were subjected to sulfonation at position six of their pyrimidine rings when treated with sodium bisulfite.1 This model was later extended to 5-methylcytosine (5mC) residue.2
In 1992, Frommer et al. described that the differing rates of converting 5mC to cytosine could be used to analyze DNA methylation patterns in genomic DNA.3 Briefly, DNA was purified from HeLa cells, placenta, liver, and sperm. A plasmid was constructed that contained the promoter and first exon of the human kininogen gene. This plasmid contained known methylation patterns and was used as a standard. The DNAs were linearized with either EcoRI or via fine needle shearing and then treated with sodium bisulfite. The samples were then dialyzed to remove any unreacted bisulfite. Dialyzed samples were dried down, the resuspended in buffer, treated with sodium hydroxide then ammonium acetate. The bisulfite reacted DNA was precipitated and resuspended in buffer. The purified, converted samples were then amplified with strand-specific primers for the human kininogen gene. The group found that it was feasible to identify specific patterns of cytosine methylation by using the discrimination in deamination reactivity of cytosine and 5-methylcytosine by bisulfite conversion.3
Now that you know how the bisulfite movement was born, we will dive into techniques to exploit this reaction. Stay tuned!
Recently I had the opportunity to meet emeritus professor Dr. Waclaw Szybalski from the University of Wisconsin- Madison, who has worked at the McArdle Laboratory for Cancer Research since 1960.
During an interview we discussed Dr. Szybalski’s amazing exit from his native Poland in 1946 following the alternating German and Soviet occupations, his education in the early days of genetic engineering, and finally the foundational work he has done in both prokaryotic and eukaryotic genetic engineering.
At age 90 Szybalski continues to maintain a laboratory with postgraduate students. At the same time (and with Promega’s assistance) he continues to support research in Poland. In May 2011, Szybalski was honored by the President of Poland with the highest order, Grand Cross of Polonia Restituta, celebrating his many scientific contributions, including: 1) establishment of the genetic basis of antibiotic resistance in bacteria;
2) multidrug therapy for bacterial pathogens and leukemia; and 3) the ability to sensitize mammalian cells to radiation. Continue reading “Synthetic Biology: Reaching Back 20 …Make that 50 Years”
When I was teaching I used to show this video to my students, both non-majors and biology majors, because it describes so beautifully what recombinant DNA is, how scientific collaboration works, and how the biotech industry was born. It’s an older video; the technology described is “old hat” for many of us in the life sciences, but it is the technology that got things going, and it’s technology that we still rely on today. Boyer and Cohen (and the film editors) do a great job telling the story of the collaboration that led to the founding of the first biotechnology company and the birth of what is now a burgeoning field.
At one time this video existed as part of a longer interview in which both Boyer and Cohen talk about the teachers and inspirations that led them into science careers. Unfortunately that is not available on YouTube, but if you can find a copy and you are working with students, I highly recommend it.
While working on a cell cycle lecture for the Education Resources web at Promega.com, I reread some classic papers describing classic cell-cycle experiments. Two of these papers describe the experiments by Murray and Kirschner showing that cyclin B synthesis and degradation are required for cycling in Xenopus oocyte extracts. When I took my first graduate-level cell biology course in 1989, these papers had just been published. I remember the instructors of the course being particularly excited about this work. (I also remember getting the events of Xenopus oocyte activation and fertilization mixed up on one of the tests for this course and realizing it about two minutes before I had to hand in my test, but I digress.)
As many of us commence our holiday festivities toasting the year’s end while earnestly drawing up personal lists of events that have shaped our lives, I would like to take a brief look at three achievements in the biological sciences—two historical and one more recent—that have struck me as nothing short of momentous in their significance. The first is the publication of a book that today continues to be an outstanding and extremely readable overview of the state of research in the genetics of animal embryology. The second is a landmark study that has brought into sharp focus the molecular mechanisms through which specific epigenetic factors modulate animal behavior. The third is the functional characterization of recBCD, a DNA-unwinding protein complex that plays a crucial role in bacterial recombination. I consider the scientists involved in each of these achievements to be pioneers— ‘podium grabbers’ if you will who have performed medal-winning science in their respective fields of expertise. Continue reading “The ‘Podium Grabbers’: Winners Who Have Medaled With Biology”