Recent advances in the generation, purification and cellular delivery of RNA have enabled development of RNA-based therapeutics for a broad array of applications. RNA therapeutics comprise a rapidly expanding category of drugs that will change the standard of care for many diseases and actualize personalized medicine. These drugs are cost effective, relatively simple to manufacture, and can target previously undruggable pathways. It is a disruptive therapeutic technology, as small biotech startups, as well as academic groups, can rapidly develop new and personalized RNA constructs. In this review we discuss general concepts of different classes of RNA-based therapeutics, including antisense oligonucleotides, aptamers, small interfering RNAs, microRNAs, and messenger RNA. Furthermore, we provide an overview of the RNA-based therapies that are currently being evaluated in clinical trials or have already received regulatory approval. The challenges and advantages associated with use of RNA-based drugs are also discussed along with various approaches for RNA delivery. In addition, we introduce a new concept of hospital-based RNA therapeutics and share our experience with establishing such a platform at Houston Methodist Hospital.

A set of instruments and specialized equipment is necessary to equip a laboratory to work with DNA. Reducing the barrier to entry for DNA manipulation should enable and encourage new labs to enter the field. We present three examples of open source/DIY technology with significantly reduced costs relative to commercial equipment. This includes a gel scanner, a horizontal PAGE gel mold, and a homogenizer for generating DNA-coated particles. The overall cost savings obtained by using open source/DIY equipment was between 50 and 90%.

Single-stranded DNA (ssDNA) oligonucleotides are useful as aptamers, hybridization probes and for emerging applications in DNA nanotechnology. Current methods to purify ssDNA require both a strand-separation step and a separate size-separation step but may still leave double-stranded DNA (dsDNA) impurities in the sample. Here, we use commercially available acrydite DNA primers to immobilize one strand of a PCR product within a polyacrylamide matrix. Electrophoresis moves the non-crosslinked DNA into the gel where the single-stranded product of desired size can be recovered. Our results show this method produces high yields of pure ssDNA.

The low-cost Open qPCR instrument can be used for different tasks in the aptamer selection process: quantification of DNA, cycle course optimization, screening, and final binding characterization. We have selected aptamers against whole Drosophila C virus (DCV) particles and recombinant epidermal growth factor receptor (EGFR). We performed systematic evolution of ligands by exponential enrichment (SELEX) using the Open qPCR to optimize each amplification step. The Open qPCR instrument identified the best aptamer candidate. The Open qPCR has the capacity to perform melt curves, and we used this function to perform thermofluorimetric analysis (TFA) to quantify target-aptamer binding. We confirmed target-aptamer binding using flow cytometry. A sandwich type luminescence bioassay based on our anti-DCV aptamer was sensitive to DCV and did not respond to a related virus, demonstrating that our selected anti-DCV aptamer can be used to specifically detect DCV.

We report the release of catalytic DNA walkers from hydrogel microparticles and the detection of those walkers by substrate-coated microparticles. This might be considered a synthetic biology analog of molecular signal release and reception. One type of particles was coated with components of a DNA one-step strand displacement (OSD) reaction to release the walker. A second type of particle was coated with substrate (or “track”) for the molecular walker. We distinguish these particle types using fluorescence barcoding: we synthesized and distinguished multiple particle types with multicolor fluorescence microscopy and automated image analysis software. This represents a step toward amplified, multiplex, and microscopically localized detection based on DNA nanotechnology.