JSPS Postdoctoral fellowship: Development of RNA-based computing for CRISPR/Cas9 with high-throughput microfluidic and next generation sequencing.
A 2 year JSPS postdoctoral position opens at LIMMS, Tokyo starting between September and November , 2017
Application deadline : March 10th, 2017
For details contact Anthony Genot ()
The Applied Microfuidic Systems lab of Prof. Teruo Fujii has been studying microfluidic devices and microfluidics since it establishment in 1999 on the campus of the Institute of Industrial Science. While the topics are ranging from basic technologies to applied research, in recent years we are putting more and more intense on the four research poles; 1) fundamental technologies in microfluidics and nanofluidics, 2) cell engineering devices, 3) microfluidic devices for deep sea in situ measurement, and 4) molecular engineering devices.
The candidate will work closely with Dr Anthony Genot, a CNRS researcher hosted in the Fujii lab and with 10 years of experience in molecular programming and DNA nanotechnology. The goal of molecular programming is to design nucleic acids to perform elaborate and useful computation in vitro and in vivo. The candidate will benefit from close interactions with a team of specialists of next generation sequencing and CRISPR/Cas9 located on the campus.
Mission:In the past few years, the CRISPR/Cas9 system has revolutionized biology by allowing programmed cleavage of DNA strands. Its versatility, robustness and programmability is such that few areas in biology have been left untouched . Cas9 and its variants have been used not only as genomic scissors, but also as spatial probes, or light-induced transcription factors. The CRISPR/Cas9 system combines a protein part (the Cas9 nuclease) and a nucleic acid part (the sgRNA which guide Cas9 to its target locus). While a lot of engineering went into improving and tweaking the protein machinery, for example to repurpose Cas9 as a programmable nickase or transcription factor, relatively less efforts were devoted to the nucleic acid part. Yet, in light of recent advances, it would be highly desirable to conditionally control the activity of Cas9 based on the presence of cognate RNAs, for example to trigger Cas9 only in particular types of cells or in response to certain sources of stress.
In parallel, the last two decades have seen the rise of Nucleic Acid (NA) technology  and, more generally molecular programming [3,4]. Exploiting the predictability of DNA base pairing, these communities have shown how to build and program molecular programs and nanostructures based on DNA and RNA, ranging from digital logic circuits, convoluted 3D shapes, oscillators, memories and neural networks. Most of those molecular programs rely on fluorescence for their readout, but the number of distinct fluorescent channels is severely limited. While Next Generation Sequencing (NGS) is a natural candidate to increase readout, it is not clear how the technology could be linked to DNA/RNA logic circuits.
In this project, we aim to combine Nucleic Acid nanotechnology and Cas9 to bring information processing to CRISPR/Cas9 system on one side, and endow molecular programs with DNA recording capabilities on the other side. Equipping Cas9 with RNA logic gates will dramatically expand its flexibility and applicability. Additionally, the DNA-editing of Cas9 will offer a drastically new way of recording the outputs of RNA-based logic circuits. Overall, it is expected that integrating logic-gated Cas9 into a wider framework of synthetic biology will bring about unprecedented benefits.
This project will be based on high-throughput technologies - such as microfluidic, combinatorial barcoding and next-generation sequencing - to harness the vast multiplexing offered by Cas9. We recently developed a general droplet microfluidic platform to simultaneously test tens of thousands of experimental conditions . Additional microfluidic systems will also be designed to combinatorially prepare and read logic circuits, building on the expertise that we accumulated in this domain.
 Jinek, Martin, et al. "A programmable dual-RNA–guided DNA endonuclease in adaptive bacterial immunity." Science 337.6096 (2012): 816-821.
 Pinheiro, Andre V., et al. "Challenges and opportunities for structural DNA nanotechnology." Nature nanotechnology 6.12 (2011): 763-772.
 Montagne, Kevin, et al. "Programming an in vitro DNA oscillator using a molecular networking strategy." Molecular systems biology 7.1 (2011): 466.
 Qian, Lulu, Erik Winfree, and Jehoshua Bruck. "Neural network computation with DNA strand displacement cascades." Nature 475.7356 (2011): 368-372.
 Genot, A. J., et al. "High-resolution mapping of bifurcations in nonlinear biochemical circuits." Nature Chemistry (2016).
Paid equivalent to the JSPS Postdoctoral position