Team:EPF Lausanne/Microfluidics

From 2014.igem.org

Next step: Designing a chip ->

Microfluidics




Microfluidics and synthetic biology


Microfluidics is an efficient tool for biological experiments. Its fields of applications go from gene regulatory network analysis to antibody screening. Several laboratory techniques can be adapted to these devices, such as DNA amplification, protein separation or cell sorting.

The chips are generally fabricated from elastomeric materials, such as polydimethylsiloxane (PDMS) and contain micron-sized channels with integrated micromechanical tools (mixer, valve, pump…). This allows massive parallelisation as well as great modularity of the experiments.

Most soluble reagents can be used, including DNA, proteins and small molecule libraries. As we focused our work on E. coli and S. cerevisiae, our experiments included on-chip culture of these species. We first used the MITOMI chip which was invented in the lab of our supervisor Pr. Maerkl. We then designed new chips that were more adapted to stress the cells by pressure, as needed to implement the final “BioPad”.


The major benefits of using microfluidic chips are:

  • Low volume required (microliter range)
  • High-throughput
  • High precision and sensitive detection
  • Low cost
  • Wide range of applications
  • Safe, enclosed environment (for more information go to the safety page)

Some examples of microfluidic experiments:

  • Transcription factors – DNA interactions
  • Protein – protein interactions
  • On-chip gene synthesis: protein expression from coding DNA
  • On-chip chemostat chambers: can be used to trace the fate of a single bacterium or to grow bacteria/yeast
  • Antibody characterisation

Why use microfluidics in our project?


As we went on with the conception of our BioPad idea, we soon had to find a solution to the question: where are we going to store our bacteria? We then came up with the idea of using a microfluidic chip as that could play the role of “container” and interface. Indeed this device fulfils most of our needs, as our support required to be cheap and easy to use. Additionally, the grid-like design of our chips, with their hundreds of chambers, was adapted to the need of having “pixels”, which would increase the precision of the readout. We also had the opportunity to use the expertise of our supervisors and to easily get in touch with the techniques of using microfluidics.

Moreover the PDMS allows the chip to be compressible and thus transmit the outer touch pressure to the bacteria in the chambers, hence we had our interface. Finally a simple but important point is the transparent colour of the chip which allows to scan fluorescence/luminescence through it.


How does it work ?


Chip sketch

  • a. Disassembled view of a microfluidic chip showing all the different components and the region where bacteria/yeasts are located
  • b. Cross section of the chip showing how a valve works: when pressure is applied in the control channel, the ceiling of the flow layer is pushed against the glass slide, which closes the flow channel
  • c. When pressure is retrieved, the ceiling elevates again, which opens the flow channel

A standard microfluidic chip is a grid of interconnected channels and chambers. It is usually composed of one or two PDMS layers placed on a glass slide. In our case we used two layers, the so called flow layer and control layer. The bacteria are enclosed between the flow layer and the glass slide. By its shape, the flow layer is responsible for the patterns of the chip. In our case, the pattern consists of several parallel rows of chambers. The control layer comes on top of the flow layer and allows to open or close valves by pressing or releasing water in the corresponding channels. Thus a mechanical pressure can be applied from the control layer on the flow layer, enabling a precise compartmentalization of the chip.

Once the chip is ready to be used, small tubings of 0.35mm diameter are plugged in the inlets of the chip (see gif below). The tubings that are plugged in the control inlets are loaded with water and enable the opening or closing of valves. The tubings that are plugged into the flow inlets are used to flow bacteria/yeast or various solutions in the chambers.


Pictures of the MITOMI chip and our Smash-Coli chip

Mitomi

MITOMI chip filled with bacteria expressing GFP

Killcoli

“Smash-coli” chip, here with
expression of RFP



Next step: Designing a chip ->

References

  1. Rockel, S., Geertz, M., & Maerkl, S. J. (2012). MITOMI: A Microfluidic Platform for In Vitro Characterization of Transcription Factor–DNA Interaction. In Gene Regulatory Networks (pp. 97-114). Humana Press.
  2. Rockel, S., Geertz, M., Hens, K., Deplancke, B., & Maerkl, S. J. (2012). iSLIM: a comprehensive approach to mapping and characterizing gene regulatory networks. Nucleic acids research, gks1323.

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