Biology subjects

Chandrasekaran, A. R.

Publications and source records attributed to Chandrasekaran, A. R..

21 records · Page 2Linked to original sources

bioRxiv2020

Programmable low-cost DNA-based platform for viral RNA detection

Viral detection is critical for controlling disease spread and progression. Recent emerging viral threats including Zika, Ebola, and the current COVID-19 outbreak highlight the cost and difficulty in responding rapidly. To address these challenges, we develop a platform for low-cost and rapid detection of viral RNA with DNA nanoswitches designed to mechanically reconfigure in response to specific viruses. Using Zika virus as a model system, we show non-enzymatic detection of viral RNA to the attomole level, with selective and multiplexed detection between related viruses and viral strains. For clinical-level sensitivity in biological fluids, we paired the assay with a sample preparation step using either RNA extraction or isothermal pre-amplification. Our assay can be performed with minimal or no lab infrastructure, and is readily adaptable to detect other viruses. We demonstrate the adaptability of our method by quickly developing and testing DNA nanoswitches for detecting a fragment of SARS-CoV-2 RNA in human saliva. Given this versatility, we expect that further development and field implementation will improve our ability to detect emergent viral threats and ultimately limit their impact.

bioengineering↗

bioRxiv2019

Exceptional biostability of paranemic crossover (PX) DNA, crossover-dependent nuclease resistance, and implications for DNA nanotechnology

Inherent nanometer-sized features and molecular recognition properties make DNA a useful material in constructing nanoscale objects, with alluring applications in biosensing and drug delivery. However, DNA can be easily degraded by nucleases present in biological fluids, posing a considerable roadblock to realizing the full potential of DNA nanotechnology for biomedical applications. Here we investigated the nuclease resistance and biostability of the multi-stranded motif called paranemic crossover (PX) DNA and discovered a remarkable and previously unreported resistance to nucleases. We show that PX DNA has more than an order of magnitude increased resistance to degradation by DNase I, serum, and urine compared to double stranded DNA. We further demonstrate that the degradation resistance decreases monotonically as DNA crossovers are removed from the structure, suggesting that frequent DNA crossovers disrupt either the binding or catalysis of nucleases or both. Further, we show using mouse and human cell lines that PX DNA does not affect cell proliferation or interfere with biological processes such as myogenesis. These results have important implications for building DNA nanostructures with enhanced biostability, either by adopting PX-based architectures or by carefully engineering crossovers. We contend that such crossover-dependent nuclease resistance could potentially be used to add "tunable biostability" to the many features of DNA nanotechnology.

bioengineering↗

bioRxiv2018

Detection of cellular microRNAs with programmable DNA nanoswitches

MicroRNAs are short non-coding regulatory RNAs that are increasingly used as disease biomarkers. Detection of microRNAs can be arduous and expensive, and often requires amplification, labeling, or radioactive probes. Here we report a single-step, non-enzymatic detection assay using conformationally responsive DNA nanoswitches. Termed miRacles (microRNA activated conditional looping of engineered switches), our assay has sub-attomole sensitivity and single-nucleotide specificity using an agarose gel electrophoresis readout. We detect cellular microRNAs from nanogram-scale RNA extracts of differentiating muscle cells, and demonstrate multiplexed detection of several microRNAs from one biological sample. We demonstrate one-hour detection without expensive equipment or reagents, making this assay a compelling alternative to qPCR and Northern blotting.\n\nSignificance statementDetection of microRNAs play a key role in biological research and medical diagnostics, and current detection methods are expensive and require sophisticated processes. We present microRNA activated conditional looping of engineered switches (miRacles), a mix-and-read strategy that is based on conformational changes of DNA nanoswitches upon binding a target microRNA. MiRacles has a sensitivity of [~]4 copies/cell and specificity of a single nucleotide, and can be performed in one hour at a fraction of the cost of traditional microRNA detection techniques. Our method can also be multiplexed to detect multiple microRNAs from one biological sample. The minimalistic miRacles assay has immediate application in biomedical research and longer term potential as a clinical tool.

bioengineering↗