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High-throughput capillary array electrophoresis for routine DNA sequencing

Objective



This project is based on the observation that the electrophoresis step in sequencing is a major bottleneck and a considerable financial burden of genome biomedical project. It is aimed at constructing a prototype capillary array sequencer with significantly increased output and decreased operation cost, as compared with slab-gel sequencers. Capillary electrophoresis combines the following advantages as compared with slab-gels:

- Capability of sustaining higher field strengths, leading to shorter separation times
- Easy automation of separation matrix replacement and sample loading
- Miniaturization, allowing a larger number of samples to be studied.

To fully benefit from this potential, however, problems remain to be solved, because capillary electrophoresis is a much more recent technique than slab-gels. In particular, the sequencing experiments performed in capillaries up to now suffered from the short lifetime of gels cast in capillaries, which renders the operating cost prohibitive, from serious band-compression problems, and from detection methods with an insufficient sensitivity/output rate. The team will overcome these limitations, by an integrated interdisciplinary approach combining competences in biology, chemistry, chemical physics and instrumentation.

The project will focus on low operation cost, high automation and robustness for routine operation, and reduce risks by exploring options on a small scale before fixing them for the full-scale prototype. It will include in particular the following innovative approaches:

- - The synthesis of new acrylamido monomers, yielding sieving matrices with longer lifetime and better resistance to hydrolysis
- - "Intelligent" new sieving matrices based on associating polymers, which can be switched from liquidlike behaviour (for automated replacement of used sieving medium) to gellike behaviour (for good separation properties)
- - A new reporter chemistry using lanthanides, for better signal/noise ratio
- - The use of pulsed-field capillary electrophoresis to increase the size range separable at high fields
- - A new technology for mass production of very-low-cost capillary arrays
- - An original detection scheme based on imaging techniques, well adapted to massively parallel acquisition

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

Institut Curie
Address
11,Rue Pierre Et Marie Curie
75005 Paris
France

Participants (5)

CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE
France
Address
Rue Georges Clémenceau Université De Paris-sud 15
91406 Orsay
MAX-PLANCK-GESELLSCHAFT ZUR FOERDERUNG DER WISSENSCHAFTEN E.V.
Germany
Address
Ihnestrasse 73
14195 Berlin
ROYAL INSTITUTE OF TECHNOLOGY
Sweden
Address
Teknikringen 30
100 44 Stockholm
THE MANCHESTER METROPOLITAN UNIVERSITY
United Kingdom
Address
Chester Street John Dalton Building
M1 5GD Manchester
THE UNIVERSITY OF MILANO
Italy
Address
Lita, Via Fratelli Cervi 93
20090 Segrate