Objective
The success of the human genome sequencing project means that DNA sequence information is one of the greatest stores of information and is now of interest to individuals. We will initiate work to construct a single-molecule DNA-sequencing device, but closely linked to this development process, we will also construct a nano-switch, based on a biological molecular motor and a moving magnetic bead. This device will provide a link between the biological world and the silicon-based microelectronics world. Such a device is likely to find longer-term uses in the development of a wide range of devices from biosensors to 'new generation' prosthetics and artificial limbs constructed with controlling computer devices. In addition, the magnetic bead attached to DNA can be used in the single-molecule DNA sequencing apparatus (to overcome coiling due to entropy) and for monitoring the movement of the DNA molecule. The success of the human genome sequencing project means that DNA sequence information is one of the greatest stores of information and is now of interest to individuals. We will initiate work to construct a single-molecule DNA-sequencing device, but closely linked to this development process, we will also construct a nano-switch, based on a biological molecular motor and a moving magnetic bead. This device will provide a link between the biological world and the silicon-based microelectronics world. Such a device is likely to find longer-term uses in the development of a wide range of devices from biosensors to 'new generation' prosthetics and artificial limbs constructed with controlling computer devices. In addition, the magnetic bead attached to DNA can be used in the single-molecule DNA sequencing apparatus (to overcome coiling due to entropy) and for monitoring the movement of the DNA molecule.
OBJECTIVES
- Site-directed mutagenesis of the motor subunit of the molecular motor to produce a motor that cannot cleave DNA;
- Determine the activity of the molecular motor (enzyme) in nano-grooves and therefore, determine the effect of large surface areas on motor activity;
- Determine the precise forces generated by the molecular motors when moving DNA, using single-molecule techniques;
- Develop single-molecule handling techniques to allow us to easily and reliably manipulate a biological, DNA-based molecular motor attached to a single DNA molecule;
- Produce highly magnetic nanoparticles for attaching to DNA in the above nano-groove;
- Investigate a variety of methods for reading DNA sequence during translocation of DNA past a sensor;
- Investigate novel DNA sequencing methods based on force microscopy;
- Develop a sensor for detecting movement of the magnetic nano-particle;
- Initiate development of an integrated system that could rapidly lead to future development of a single-molecule DNA sequencing device.
DESCRIPTION OF WORK
We have available a number of unique DNA-based molecular motors that differ from other DNA-based motors in that they do not track along DNA in a linear fashion, but move the DNA toward the bound motor/DNA complex. As such they are nano-actuators capable of moving any object attached to the end of the DNA toward the motor/DNA complex. One example of such a motor is a type I restriction-modification enzyme, which could under certain circumstances cleave the DNA. To remove this possibility we will alter the amino acids required for this function using site-directed mutagenesis. We will then assay activity of a variety of motors, at low concentration, in silicon microwells and channels to determine whether silicon surfaces interfere with motor activity. Single molecule handling techniques will be used to precisely determine the force generated by the motor molecules. In addition, force microscopy techniques will be explored as a method for detecting single base-pair mis-matches in DNA sequences, providing an alternative technology for reading important DNA information. A variety of methods will be used to determine the easiest and most reliable method for localising a single DNA molecule in a sub-micron silicon groove. In parallel to the above work, we will develop novel nano-scale magnetic particles and investigate their magnetic properties using a variety of techniques including micro-SQUID, Hall-effect sensors and Magnetic Force Microscopy (MFM). This will lead to the development of a nano-sensor for detecting magnetic field changes. Finally, we will investigate the potential for fluorescence resonance energy transfer (FRET), scanning tunnelling microscopy (STM) and scanning near-field optical microscope (SNOM) with fluorophores located at the end of nanotubes will be used to detect the movement of DNA past the sensor. This will allow us to evaluate each technique as a potential means for reading the DNA sequence on a single molecule.
Fields of science (EuroSciVoc)
CORDIS classifies projects with EuroSciVoc, a multilingual taxonomy of fields of science, through a semi-automatic process based on NLP techniques.
CORDIS classifies projects with EuroSciVoc, a multilingual taxonomy of fields of science, through a semi-automatic process based on NLP techniques.
- natural sciencesbiological sciencesgeneticsDNA
- natural sciencesphysical sciencesopticsmicroscopy
- natural sciencesphysical scienceselectromagnetism and electronicsmicroelectronics
- natural scienceschemical sciencesinorganic chemistrymetalloids
- natural sciencesbiological sciencesbiochemistrybiomoleculesproteinsenzymes
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Topic(s)
Call for proposal
Data not availableFunding Scheme
CSC - Cost-sharing contractsCoordinator
PO1 2UP PORTSMOUTH
United Kingdom