BioNano-Switch EC Project
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Management Report No 2 (Summary of Progress)
This Project is now in its final year and significant progress
has been made toward the original concept. The aim was to produce three modules
of a prototype device that would act as a generic biosensor, each with its own
independent capability, but each is also intrinsically linked in the final,
proposed, device (Figure 1). This modular approach was seen as an ideal example
of Synthetic Biology, where biomaterials would be used to construct an
artificial device using engineering principles to guide the construction. One
such engineering approach is to demonstrate that such modules could be linked
together in a functional device. We have reached a stage within the Project
where we believe a commercial device is a realistic possibility.
As we approach the final stages of this Project we can
report significant progress toward the above Objectives:
- The recombinant DNA work, at both Portsmouth and IMIC,
has produced molecular motor proteins linked, via protease cleavage sites,
to GST that will enable immobilisation. In addition, a biotinylated
protease that can be linked to the biotinylated antibodies (commercially
available in ELISA kits) via a biotin‑streptavidin‑biotin-protease linkage
are now available. We have, therefore, made significant progress toward a
commercial device, using the Molecular Amplifier concept. In addition, we
now have all of the components for Module 1 and 2 and we are also testing
methods for integration of these Modules within the proposed device.
- Aptamers, as an alternative to antibodies for Module
1, have been produced and characterised together with a model system (a
thrombin‑binding aptamer). Surface attachment has been investigated, but
release and re-attachment remains a problem. Novel aptamers have been
raised to the HsdR subunit of the motor system EcoR124I and the PrescissionTM Protease.
- Work on the construction of a GST-(presc.prot)‑FtsK,
as an alternative molecular motor for translocation of DNA, has recently
been successful. To enable this, we instigated a more extended
collaboration between IMIC and UoP (via an appointment of a graduate
student, from Portsmouth, based in the IMIC laboratories), which has
contributed to the success of this work.
Investigation of other motors has also taken place, including helicases and
topoisomerases; a novel method of site
specific tethering to DNA of the Rad54 motor was developed. This allows the
Rad54 motor, as well as other translocating motors, to efficiently create
DNA loops. Successful analysis of helicases in the Magnetic Tweezer system
has led to the award of a new Research Grant (under the NanoSci-E+ scheme)
to UoP, ENS and INESC‑MN (together with an additional Partner at TUDresden)
to study helicases from Plasmodium as potential targets for anti-malarial
drug targets – an exciting development from the BioNano-Switch Project.
magnetoresistive sensor system, based on Spin Valves and MgO Magnetic Tunnel
Junctions, have been integrated with in‑chip current lines at the sensor level
and microfabricated in glass substrates. The focus/placement of microspheres
flowing in PDMS based microchannels bonded to the chip, the stretch and magnetic
excitation of magnetically labelled biomolecule (dsDNA bonded to a gold pad
above the sensor) are both achieved with in-chip structures allowing further
Work on the Hall Effects Sensors was focused on the reliability of the FEBID
process and allowed improvement of the field resolution below 1 µT/Hz0.5,
which, according to calculations, is more then sufficient for bead detection.
Alternatively, CoPt Hall sensors with field resolution down to 1 nT/Hz0.5
were investigated for applications where spatial resolution is important, but
these sensors need to be properly biased, as the working field is limited to a
few mT, similar to magnetoresistive sensors.