Team:Austin Texas/human practices

ELLA

• Content - some data, some of the discussions/outreach we had with people.

Caffeinated Coli

Ella collecting coffee sample from Austin Java

[http://www.google.com/maps/d/viewer?mid=zlUDhks6KHxI.ketDvlvg6zis Click here for an interactive map of coffee shops we visited!]

Jordan discussing our project with Matt Bolick, co-owner of Flat Track Coffee

Previously, the UT Austin iGEM team had used the caffeinated coli to measure the caffeine content in beverages such as soda and energy drinks. The results showed that these engineered E. coli were very accurate at detecting the levels of caffeine in these formula-based drinks. We wondered how well they would do with something different, something that the vast majority of people in the city of Austin drink on a regular basis, coffee.

The [http://www.ibuyaustin.com Austin Independent Business Alliances'] slogan of [http://www.keepaustinweird.com "Keep Austin Weird,"] is a widely shared sentiment of Austinites, and emphasizes the need to promote local businesses. Thus, we thought it would be a great community outreach project to work with local Austin coffee shops, and do our part to "Keep Austin Weird!" Jordan, a senior member of the group, took charge and assembled a map of about 40 local coffee shops in Austin. The team took a Saturday and split up to visit dozens of coffee shops, collecting samples of their house coffee. We were met with much enthusiasm by the coffee shop owners and employees, whose interest had been piqued when we described our experiment, often resulting in discussions of science, synthetic biology, and coffee.

Back at the lab, the team members got to work. Senior members divided their attention to mentor the new members in the techniques necessary to start cultures, work with the lab equipment, and other general laboratory safety.

The beginning of something great! Special thanks to Razan Alnahhas (second row, right) for mentoring and to Mike Hammerling (not pictured) and Ashley Kessel (second row, left) for assistance in sample collection.

Measuring Caffeine Content

Cultures of knockout strain grown with collected coffee samples.

Standard growth curve of knockout strain grown with different amounts of caffeine.

The E. coli we used are an engineered strain, previously made by the 2012 UT Austin iGEM team. This strain has had its normal guanine synthesis pathway knocked out, but contains a plasmid with a set of genes that enables the organism to synthesize guanine from xanthine and many of its derivatives, including caffeine. Without caffeine (or other xanthines), the strain cannot grow. When caffeine is added, the strain can demethylate the caffeine moleculesm yielding xanthine, and from there the E. coli can synthesize guanine. It then grows normally until the caffeine is completely metabolized, at which point the cells can no longer synthesize guanine, and cease to grow. Thus, by looking at the relative growth of the strain with different samples of coffee and comparing it to a standard curve of growth in solutions with known amounts of caffeine, we could reliably measure the amount of caffeine in the coffee.

The protocol was simple: We provided our Caffeinated coli with a diluted sample of coffee we had acquired from the various shops, and then compared the relative growth of each culture by measuring the absorbance at 600 nm (OD600 for short). The simplicity of the project was the key its success. It provided a glimpse into the exciting new world of synthetic biology for the new members and allowed them to learn vital skills that were used in day to day research during the summer. Our results can be seen in Figure 1, below.

Figure 1. Relative levels of caffeine in house coffee samples from around Austin. Data depict the amount of caffeine in the coffee relative to the average caffeine content, as determined using the caffeinated coli.

SXSW Create

A couple of weeks later, we presented this data set at the SXSW festival. As part of this outreach, we explained our project and synthetic biology to a wide range of people. Many people who came by had little to no background in science or technology, and even most of the ones who did had not heard about synthetic biology. By discussing our experiments with bacteria and coffee, an interesting and easy to understand application, we were able to explain what synthetic biology is and how it can be used in a beneficial way. Occasionally, these conversations would branch out into wider topics of science and technology.

People were particularly interested in the different types of coffee that yielded strange results. For example, it appeared that there was slightly more caffeine in the Epochs' Light Roast, than in the Dark roast, which seems to contradict the modern myth that dark roast coffee has more caffeine. There was much debate sparked when people started to make speculations on why that was so. Some people claimed that longer roasting destroyed the caffeine (and swore by cold-press coffee!), some people guessed that perhaps the dark roast sample had less coffee beans in it when it was brewed. Nevertheless, after much research by the team after the event, it was concluded that [http://en.wikipedia.org/wiki/Coffee_roasting coffee roasting] and [http://en.wikipedia.org/wiki/Coffee_preparation#Brewing coffee brewing] are largely variable processes, and caffeine content can hugely vary from batch to batch of beans. For more information about light roast vs. dark roast see this article at [http://www.scribblerscoffee.com/coffees_caffeine.htm Scribblers Coffee], which discusses the nuances between the two!

People also were extremely curious as to why the Wright Bros. Brew & Brew sample had a significantly more amount of caffeine. This data point compelled us to speak to one of the owners of Wright Bros. Brew & Brew, to determine how they had created such a highly caffeinated beverage. The owners concluded, after discussing our project and the coffee brewing process, that the specific pot of coffee was probably just not fully drained, i.e. there was less water in the batch than normal, resulting in a higher caffeine/volume content. However, similar conversations were had between the entire team and the various coffee shops and baristas during the collection and presentation processes.

Coffee bean counting contest

To involve everyone attending the event and raise more awareness for the iGEM team, we held a contest in which patrons attending the event could enter to win an E. coli plush upon following us on Twitter.

Special thanks to Karen Ingram for giving us this amazing opportunity to participate at SXSW Create 2014!

Photographs