PFI:AIR - TT: Dielectrophoretic Fractionation of DNA for Next Generation Sequencing

Project: Research project

Project Details

Description

Overview:
This Accelerating Innovation Technology Translation project aims in developing a versatile tool for nucleic acid fractionation for the ever increasing field ofnext generation sequencing (NGS). The highly dynamic and fast growing field of NGS requires the isolation of suitable size fractions of nucleic acids as a crucial prerequisite for library construction. Currently available methods are cumbersome and not flexible enough to keep up with the rapid technical advancements in this field. Here, we propose the development of a microfluidic sorter to overcome the current bottleneck in NGS applications by exploring the proof-of-concept for a DNA fractionator based on dielectrophoresis (DEP).
Depending on the platform NGS requires specific DNA lengths in the range of 200bp to a few kbp for efficient sequencing. The currently available methodology on the market to prepare desired size fractions has limitations due to cumbersome sample preparation requirements and the restricted applicability for a specific size range only. In this proposal, we will develop a proof-of-principle sorter that will allow fractionating DNA in desired size ranges and importantly will be applicable to the entire spectrum of DNA lengths currently suitable for NGS. We will also investigate the application of the novel microfluidic sorter for RNA fractionation by length. The microfluidic approach will further provide a cost effective device requiring minimum DNA and RNA sample, recover size fractions in solution and thus allow their direct application on NGS platforms and importantly will allow adapting DNA and RNA size sorting into tunable size fractions on a single device. We thus expect to develop a versatile new tool with wide applicability to current and future NGS platforms.

Keywords: Next generation sequencing, microfluidics, lab-on-a-chip, insulator-based dielectrophoresis, nucleic acid, DNA, RNA, size fractionation, women in chemistry

Intellectual Merit:
Undoubtedly, genome sequencing is among the most powerful current bioanalytical techniques. The advances in next generation sequencing allowed deciphering a complete human genome within one week compared to the first human genome, which took more than 10 years (and about 3 billion US dollars). A single NGS run still costs in the order of $5k-10k not including sample preparation. A bottleneck in NGS is the various size fractions and cumbersome sample preparation techniques for each specific platform. With the use of our novel dielectrophoretic technology we overcome these limitations and aim in developing a versatile device capable of delivering DNA in size ranges suitable for all current and future NGS platforms. Additionally, the development of our proof of principle device will contribute to the field of biomolecular DEP and provide valuable knowledge on this relatively new migration mechanism for biomolecules and specifically DNA.

Broader Impact: Genome sequencing is important for deciphering biological pathways leading to malfunction and disease. Facilitating genome sequencing with our proposed approach will thus have important contributions in many fields of biology and medicine, including disease combat, drug development and diagnostics. A versatile sample preparation tool will further allow reducing costs of genome sequencing making NGS analysis applicable to broader fields in science and eventually facilitate personalized medicine due to a broad availability of NGS.
The project will further be tied to the mentoring activities for female students of the PIs currently active Career Award. These activities are bundled in the REACH (Research and Activity Platform for Female Chemists) program at the Department of Chemistry and Biochemistry at Arizona State University. We aim in involving a female undergraduate student in the proposal by providing an undergraduate research opportunity for one year. Furthermore, a female scientist from Qiagen will be involved in the REACH mentoring activities providing another valuable industry component to the mentoring program.

Description

The underlying Accelerating Innovation Technology Translation project aims in developing a versatile tool for nucleic acid fractionation for the ever increasing field of next generation sequencing (NGS). The highly dynamic and fast growing field of NGS requires the isolation of suitable size fractions of nucleic acids as a crucial prerequisite for library construction. Currently available methods are cumbersome and not flexible enough to keep up with the rapid technical advancements in this field. We are currently developing a proof-of-principle sorter that will allow fractionating DNA in desired size ranges and importantly will be applicable to the entire spectrum of DNA lengths currently suitable for NGS. The microfluidic approach will further provide a cost effective device requiring minimum nucleic acid sample, recover size fractions in solution and thus allow their direct application on NGS platforms and importantly will allow adapting nucleic acid size sorting into tunable size fractions on a single device. We thus expect to develop a versatile new tool with wide applicability to current and future NGS platforms.

Benefit for awarded PFI:AIR project
Our recent discovery showing nDEP in insulator-based dielectrophoretic manipulation of DNA has opened a new perspective for the PFI:AIR award. Based on this nDEP behavior it becomes possible to investigate the fractionation of DNA similar to what we have shown in preliminary work with mitochondria [2] and protein nanocrystals [1]. This was not planned during the original submission. The REU opportunity will this allow to exploit nDEP for DNA fractionation. This topic is ideally suited for a summer research project, since the Ros lab has substantial experience with this fractionation under nDEP conditions both experimentally and in simulation.

Description

With this travel extension opportunity, we secure the attendance of Dr. Paul V. Jones at the PFI:AIR-TT grantee meeting 2016. This is a unique opportunity for the project PI, Dr. Jones and collaborators since the conference will provide networking and discussion options with other PFI:AIR-TT grantees. We expect to learn about successful industry-academia collaborations and also potential marketing and commercialization strategies for the technology development of our currently funded proposal entitled PFI-AIR-TT: Dielectrophoretic Fractionation of DNA for Next Generation Sequencing. We also aim to get in touch with SBIR grantees to seek advice for future project developments and expect to present our nucleic acid fractionation device for next generation sequencing to a broad audience.
StatusFinished
Effective start/end date8/15/1412/31/17

Funding

  • National Science Foundation (NSF): $249,045.00

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