Bryan Hernandez

that's me

== Bio ==

• I am currently a UROP in the Endy Lab working with Jason Kelly.
• MIT Class of 2009; Majoring in Mathematics and Biological Engineering.
• NorCal=home

Projects

Sortostat

• We are currently using strain MC4100 E.Coli capable of expressing CFP and YFP (Cyan Fluorescent Protein and Yellow Fluorescent Protein, contained on plasmid pSB1A2 ) as these are easily distinguished between visually thereby aiding in our descriminitive assortment without relying on chemical selection. In this way, it is less likely for cells to avoid being sorted by a genetic mutation as one might expect using an antibiotic.
• Chemostat Theory
• Sortostat/Experiments
• Sortostat/Growth Tests

...

Project Goals

• UROP Proposal

• Sortostat

1. Debug a microfluidic chemostat (Sortostat) to improve the time-varying specific selection of cell populations.
   • Currently troubleshooting problems: i) Cell Death after 3-4 days (presumably due to oxygen depletion,) and ii) Inaccurate cell counts due to poor image processing.
2. Evaluate the response of populations of E.Coli cells containing engineered genetic circuits (http://parts.mit.edu) to particular selective pressures using the Sortostat.

Device Longevity Characterization

It is known that mutations are more likely to occur with higher cell division rates. Everytime a cell divides it runs the risk of making a mistake in the replication process and creating a mutant cell. Evolution is largely in debt to this phenomonon; without mutants an organism's genome would be largely static. However, beneficial this might be to the survival of a cell species, it poses a problem for our genetically engineered cells. If a cell divides and mutates in the process it could "break" our engineered genetic device. The cell will likely go on living, however, it will cease to perform its intended function. This is an unfortunate reality of biological engineering, and, as such, we must learn the characteristics of our devices and their liklihoods of "breaking."

In order to obtain a quantitative measure of how "fast" our devices break we will first need a controlled environment. By using a chemostat we can control the rate at which cells divide enabling us to calculate the number of cell divisions that have occured over some time. Note that number of divisions is but one variable that can be used to characterize the longevity of a device. One might imagine testing other variables such as temperature or cell cycle position for example, both of which likely have an effect on the "breaking rate."

Detecting when a cell's device has broken can be a challenging in it of itself. We are using a counter selectable marker that will tell us when the cell's device ceases to perform its function. This marker will allow cells to grow on a plate when the cell has broken. In other words, by taking a sample of the cells from the chemostat and plating them with the counter selectable marker present, only those whose devices have broken will grow. If all of the cell's devices are fully functional, however, we expect to see no growth on the plate. pKSS is capable of providing this type of device in cells.