Ultrafast and highly multiplexed genetic identification using 2D-PCR amplification of padlock probes – FASTGENE-HM

The purpose of this project is to develop a process and a genetic identification platform enabling to detect in less than 15 minutes the presence and the concentration of one or several pathogen agents in a sample among 140 potential targets.
In many situations, it is preferable to be able to detect the presence of certain microorganisms in a given environment as fast as possible. For the armed forces deployed for a military operation, it can be vital to be able to detect in few minutes the presence in air sample of pathogen microorganisms dispersed by the aggressor. For civil applications, needs are similar for the rapid diagnostic of the presence of pathogen agents in a biological sample and the determination of an antibiotic or antiviral resistance profile.
To address these needs, the exploitation of the genetic information contained in these samples is the most promising method. One can obtain this information with two approaches:
- An analytical approach carried out by next generation sequencing, that enable to access to a random sample of this information, which is then bioinformatically analyzed. But the current generation remains too expensive, too slow and not robust enough.
- A targeted approach with the direct research of the presence of one or several genetic sequences by PCR or similar amplification methods. To be able to address the needs of certain applications, this approach must be improved relatively to the state of the art to enable the simultaneous identification of a hundred genetic sequences in few minutes. It is also crucial that this approach is quantitative and enables the determination of the respective concentrations of the present targeted sequences.
In a recent project supported by the ASTRID program, ELVESYS developed the world fastest PCR technology thanks to its expertise in microfluidics. The objective of this ASTRID MATURATION project is to extend the performances of this technique to obtain an ultrafast, quantitative and highly multiplex process and PCR system. For that purpose, we plan to use padlock probes and thermally crosslinking gels that will fully take advantage of the internal structure of our microfluidic chip and our thermalisation rapidity to improve the multiplexing capability. The simultaneous controle of these technologies within a system will enable to develop the most accomplished device for the ultra-fast detection of pathogen agents, for civil or military applications.
In terms of performance, the final objective is to be able to measure in a DNA sample, in a single reaction and in less than 15 minutes, the respective concentrations of all the sequences, among 140 targeted sequences, whose concentration is superior to 1/10000e of the most represented targeted sequence.
These performances will be demonstrated with 2 model applications: 1) a military application demonstrating the fast identification capacity of simulating agents in environmental samples, 2) a civil application with the fast diagnostic of pulmonary infections and the associated antibiotic profile.