Class Notes: Week 7

Coliformers from Melbourne's iGEM 2007 team. (Courtesy of Phillip Dodson. Used with permission.)

Abstraction hierarchy for a biological system.

Abstraction hierarchy example: arsenic detector.

Diagram mapping the arsenic detector to the system and device layers. (Figure by MIT OpenCourseWare.)

After just 5 minutes we'll see the device level system diagrams that you've drawn for this system and discuss any outstanding questions or concerns.

And finally: Polkadorks

Let's try a more dynamic system. The Polkadorks 2004 IAP iGEM team wanted their engineered cells to "form, diffuse, and form again in random areas on the plate. Our system should thus form time-varying patterns based on local random time-varying symmetry breaking." Check out the Polkadorks animation.

Animation of the Polkadorks system. (Courtesy of Ziyan Chu, Roshan Kumar, Stephen Lee, and Joe Levine. Used with permission.)

 

• As a class we'll
  • describe the system in plain language, then
  • list the devices needed to implement the system
• Then as a team you'll have 10 minutes to draw a device-level system diagram.
• Finally, as a class, we'll look at the timing diagram that the 2004 IAP team wrote.

Timing diagram from Polkadorks. (Courtesy of Ziyan Chu, Roshan Kumar, Stephen Lee, and Joe Levine. Used with permission.)

You'll see that the devices (or the named connections between these devices) are listed on a y-axis and time is shown on an x-axis. The timing diagram indicates the timing for operation of each device or wire, including the persistence of each device's signal through time—shown as PS (protein synthesis) and PD (protein degradation). You and your team should take 10 minutes to make a timing diagram for the Polkadork's devices that you have wired together. You can keep a running list of any uncertainties. After 10 minutes of work, we'll have each team report back to the group to see how we did and what questions still remain.

Part 2: Get Busy!

Start applying these ideas to your team's project. You can work on a device list, a device-level system diagram and a timing diagram if you're ready. Be sure to keep your project notebook up to date and help keep each other great.

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Lecture 13: Abstraction in Action Part 2

We've been working hard this week to move between the System and Device levels of an abstraction hierarchy and today we'll drop down one more level to think about the parts that make up a device. The device we'll consider is switch made from RNA, but first we'll look at a "classic" device, namely an inverter, and the parts that make it up.

Challenge: Abstraction in Action: Devices to Parts

Part 1: Four-part Inverter

Recall the Eau d'coli project from the 2006 MIT iGEM team. Their goal was to replace the nasty smell of bacteria with wintergreen smell during log phase growth and banana smell during stationary phase growth. In fact some of you added the banana smell generator (BSG) to bacterial cells yourself thanks to the Foo Camper's Guide to BioEngineering that we tried way back in Week 4 of this term. We also talked about the engineering ideas behind the Eau d'coli project way WAY back in Week 1. Today we'll focus in on one device within the system, namely the inverter that was used upstream of the wintergreen generating device (WGD), turning it off during stationary phase.

Abstraction hierarchy example: Eau d'coli.

The abstraction hierarchy shown above zooms in on the inverter part that reverses the signal from the stationary phase promoter. This turns the wintergreen generating device off in stationary phase and on in log phase. The inverter part that's shown is a permutation of a natural gene that encodes the tetracycline repressor protein.

Eau d'coli inverter part.

Within this gene, we can define 4 "parts:" a promoter, a ribosome binding site (RBS), an open reading frame (ORF) that encodes the tet repressor protein, and a transcriptional terminator so the RNA polymerase transcribing the gene doesn't continue down to the next sequences downstream. The promoter is repressed by the tetR protein itself, allowing for a simple rearrangement that makes a useful inverter device. You can imagine a family of 4-part inverters that might be made for every repressor protein we know (lac repressor, lambda repressor, etc). The number of these proteins is finite, however, and there will almost certainly be a time when a transcription-based device will not be optimal, so for the next part of today's challenge you are asked to make an RNA-based switch. This challenge will draw upon your understanding of basic biology, your ability to find resources, and your ability to work as a team. Good luck!

Part 2: Let's Make a Switch

Working in your project teams, develop a design for a genetically encoded switch that works through the expression of a regulatory RNA. Your team's design should include (a) a high level system diagram, (b) a full list of devices and parts, (c) a source for all the necessary DNA parts (be it research lab or synthesis company), (d) a plan for testing the most important components of your switch, and (e) the final cost associated with building and testing your device. You have 45 minutes. Your team's budget for this project is $1000. The winning team will have the most versatile RNA-switch that costs the least to construct and test.

• Note: This activity features an "All questions answered" work environment. Ask lots of questions.
• Hint: Your DNA synthesis budget may not be large enough to pay for synthesis of all the parts needed to make the switch.
• Hint: Your team may not have enough time to design everything needed to make the RNA-based switch.
• Hint: Spend 2 minutes right now thinking about all the things that need to come together over the next 45' for your team to be successful.
• Question: How will you check that everybody on your team understands what is going on?

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