CAREER: DNA Analysis Based on Dielectrophoresis Project Summary The increasing complexity of current analytical challenges demands novel and efficient separation and purification techniques. For example, the elucidation of molecular level biological complexity requires powerful separation and purification techniques, and these are often the limiting component in biochemical research such as in biomarker discovery, clinical diagnosis or single cell analysis. The separation and purification of novel macromolecular structures such as artificial DNA nano-assemblies is further essential for their successful nano-technological applications, such as for DNA computing, in photonic devices or in targeted diagnostics. In this proposal, a novel parameter will be exploited for the analysis of DNA, namely the polarizability. In an inhomogenous electric field the polarization of DNA gives rise to dielectrophoretic transport or concentration, which is proposed as a novel selection criterion for DNA analysis. To date however, the polarization mechanism of DNA remains only little understood, which hampers the use of dielectrophoresis (DEP) for applications in separation and analysis. The proposal thus aims at a quantitative study of the polarizability for a wide variety of DNA. The necessary electric field gradients probing the polarizability of DNA will be created on insulator-based dielectrophoretic devices integrated into microenvironments. Moreover, concomitantly occurring transport mechanism will be investigated to reveal their interplay with DNA DEP with the ultimate goal to optimize DNA analysis. Based on this knowledge, analytical applications ranging from DNA nanotechnology, quality control of DNA vaccines to DNA-based diagnostics are proposed. Intellectual Merit Dielectrophoresis (DEP) is capable of probing the polarizability of DNA, which is an intrinsic molecular property. The lack of understanding the polarizability of DNA has, to date, hampered the use of DEP in analytical applications. In this project, the polarizability and dielectrophoretic behavior of DNA will be quantitatively studied in a broad size range, for various topoisomers and artificial DNA nanostructures such as DNA origami. Based on these systematic and quantitative studies, significant contributions to elucidate the origin of DNA polarizability are expected. Our systematic studies will lay the foundation for novel analytical techniques with application in nanotechnology or diagnostics. The employed microfluidic platforms further provide miniaturized and fast analysis of DNA by DEP, and will allow the analysis of minute samples in the range of picoliter to nanoliter suitable for the analysis of DNA in small cell ensembles or even single cells. Per se, DEP of DNA represents a novel selectivity factor, which can be employed to manipulate nanomaterials of synthetic or biological origin. Broader Impact 1) Scientific: A detailed understanding of the polarizability and the dielectrophoretic behavior of DNA allows exploiting subtle differences of various DNA and thus extends current analytical technology with a new selectivity parameter. Analysis based on DEP will provide a novel tool, which will eventually find widespread application in analytics of natural and artificial DNA, for DNA based vaccines and diagnostics. This novel selectivity factor has also the potential to be coupled with other established techniques to create multidimensional separation approaches and be applied for the analysis of a broad range of biomolecules. 2) Broadening Participation of Women in Chemistry: Within this project, a mentoring plan for female undergraduate and graduate chemistry students will be launched in the Chemistry and Biochemistry Department at ASU. The proposed activities aim in encouragement and promotion of women, which are still underrepresented in chemistry at higher career stages. The plan includes individual mentoring activities as well as general activities for female undergraduate students including research opportunities related to the intellectual merit of this proposal. At the graduate level, the plan specifically intensifies this mentoring activities to ameliorate communication, scientific presentation and negotiation skills as well as networking opportunities, PhD progress and career development. This mentoring plan is well supported by the Department of Chemistry and Biochemistry as well as other NSF funded projects related to broaden the participation of women in science, technology, engineering and mathematics. This mentoring program will be provided for more than 100 female chemistry majors and ~60 female graduate students and thus a broad impact on gender equity is expected. Introduction and Specific Aims The increasing complexity of current analytical challenges demands novel and efficient separation and purification techniques. For example, the elucidation of molecular level biological complexity requires powerful separation and purification techniques, and is often the limiting component in biochemical research such as in biomarker discovery, clinical diagnosis or single cell analysis. The separation and purification of novel macromolecular structures such as artificial DNA nano-assemblies is further essential for their successful nanotechnological applications. The PIs active Career Award proposes to study the dielectrophoretic properties of DNA as well as DNA-assemblies with the ultimate goal to add dielectrophoresis (DEP) as a novel separation criterion to the portfolio of analytical and separation scientists. While the Career Award originally suggested investigating naturally occurring and artificial DNA in a broad variety, the proposed collaborative activity with the ERC-funded collaborator Prof. Schmidt at Georg-August-University, Goettingen, Germany, will translate the dielectrophoretic separation problem to single-walled carbon nanotubes (SWNTs) and their assemblies with ssDNA. These SWNT-ssDNA assemblies will be employed by the European host as an in vivo sensor of nuclear mRNA to investigate gene activation related to mechanical stress in selected cell lines. The major goals of the proposed collaborative activity are: i) Characterize the dielectrophoretic properties of the SWNTs employed in the ERC-funded partner project: We will investigate the dielectrophoretic properties of SWNTs with respect to length, metallic, quasi-metallic and semiconductor properties as well as with and without ssDNA wrapping in tailored microfluidic devices. ii) Design of size-based fractionation devices for SWNT-ssDNA assemblies: Based on the DEP properties revealed in aim 1, we will design microfluidic DEP-based fractionation devices for micro-quantitative SWNT-ssDNA recovery in size ranges suitable for in vivo sensing (ideally down to size fractions of ~100nm). iii) Application of the sorted SWNT-ssDNA assemblies for in vivo sensing: Fractionated SWNT-ssDNA nanoassemblies obtained from aim 2 will be used for in vivo sensing of mechanical stress in the Schmidt lab on selected cell lines.
|Effective start/end date||4/15/12 → 9/30/18|
- National Science Foundation (NSF): $513,936.00
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