On-chip Massively-parallel Electronic Detection of DNA Binding Events
Abstract
The analysis of nucleic acid (DNA and RNA) sequences is a fundamental capability of the modernage. In medicine, it enables the diagnosis of infection by pathogenic microorganisms, the identification ofgenetic variation and disorders, and the tracking of cancer progression. With the development of prototypesfor large-scale DNA-based information storage, the ability to analyze DNA has taken on a technologicalsignificance. Nucleic acid analysis can be divided into two sub-capabilities: sequencing and detection. Insequencing, the detailed letter-by-letter base composition of a nucleic acid is determined. In detection, thepresence or absence of a particular fixed-length sequence is determined. Sequencing is necessary in cases where the variation in a population of DNA or RNA strands is unknown, e.g. when one is trying to understand the detailed evolutionary history of a virus or the heterogeneity of a cancer. Since 1977, sequencing technology has progressed through a number of technological generations which have dramatically dropped the price and increased the speed of sequencing. In the last 20 years the cost of sequencing a human genome has fallen by a factor of 100,000 and the time required has dropped by roughly a factor of 300. Capital costs for equipment have plunged so that the price of the sequencing device (in the case of Oxford Nanopore???s Minion sequencer) is now less than that of the consumables used in the device ($500-$900 per ???cell???). Further, the most advanced sequencers now have the same form factor as most consumer electronic devices???about the same size as a cell phone with a USB-computer interface. In contrast to sequencing, detection of DNA sequences is sufficient in situations where the relevant sequences are already known, as in the case when one is trying simply to identify a pathogen of known strain, measure the response or recurrence of a cancer during treatment, or distinguish between artificial DNA barcodes used to label cellular or molecular libraries. Sequencing can be used for all of these tasks, but detection of DNA sequences may perform the task more quickly and cheaply. We propose to combine reconfigurable DNA devices, with an ability to position these devices on semiconductor surfaces, to create an electronic detection platform whose price (for consumables) will be 100-fold less than that of DNAsequencing, and which will be cell phone-sized for portability. Specifically, we will create ???Venus flytraps??? from DNA origami, whose top and bottom lids will carry probes designed to capture a single-stranded target DNA of interest. Upon capture of the target, the flytrap will close, creating a gross (100 nanometers or more) movement of the top lid. Detection of this conformational change will be explored using two methods: (1) square-wave voltammetry of flytraps labelled with redox-active signal molecules on gold electrodes, and (2) the label-free capacitance response of graphene field-effect devices. Because the flytrap lid will be 200-fold larger than the typical target DNA, and may carry 200 times more signaling molecules, the response will be greatly amplified and may allow single molecule sensitivity. Depending on detection method, signal will be maximized either by optimizing flytrap geometry to bring signal molecules within a couple nanometers of the surface, or adding polyethylene glycol to overcome Debye screening at the surface. Signal-to-noise, gain, specificity, and binding kinetics will be studied as a function of flytrap design. Because applications from tumor detection to DNA storage may require the detection of double-stranded DNA, we will explore flytraps that feature more sophisticated strand-displacement or CRISPR/dCas9-based binding mechanisms which do not require target DNA to be single-stranded. Fabrication challenges will also be addressed: for single-molecule measurements costly e-beam fabrication will be replaced with scalable UV photolithography.
Document Details
- Document Type
- DoD Grant Award
- Publication Date
- Sep 04, 2018
- Source ID
- N000141812649
Entities
People
- Paul W. K. Rothemund
Organizations
- California Institute of Technology
- Office of Naval Research
- United States Navy