Tech Blog: UA College of Science Tech Provides Guarantee in RNA Amplification

As a major biological macromolecule, ribonucleic acid (RNA) is essential for all forms of life [1]. In regards to infectious diseases, RNA is capable of carrying the viral genetic information [1]. In order to access and analyze this genetic information, researchers must use RNA amplification methods, which “involve a series of intricate procedures to amplify genetic signals from minute quantities of starting materials” [2].

Unfortunately, the quality of RNA present can be low due to the tiny amount of RNA initially present in many clinical samples or damage caused while samples are being prepared or stored. Often times current amplification technologies, which require numerous steps, expertise and specific facilities, will report no detectable RNA because of these challenges.

UA Ecology and Evolutionary Biology Department Head and professor Michael Worobey, Ph.D., felt the impact of this unreliable technology in his research to identify samples that may contain HIV from a time well before the virus was discovered.

Worobey has dedicated his research to understanding “the origins, emergence, and control of pathogens,” specifically retroviruses and RNA viruses like influenza and HIV [3]. Because his research focuses so heavily on the study of RNA, Worobey needed an alternative for the cumbersome and unreliable existing amplification methods.

He describes the new technology, which was developed in his lab in conjunction with UA Senior Research Specialist Thomas Watts, as a “very sensitive method for retrieving genetic material from heavily damaged samples.”

In his 2016 Nature publication, Worobey detailed how his technology aided RNA recovery as it related to studying the emergence of HIV in the United States saying, “recovered RNA was generally below the limits of quantification and initial attempts at amplification of reverse-transcribed viral RNA failed consistently…This led us to design an RNA ‘jackhammering’ approach to greatly increase both the ability to detect viral RNA-positive samples and to recover complete genomic HIV-1 sequences from them” [4]. This ultra-sensitive technique breaks the problem down into tiny pieces and rebuilds the complete genome from those pieces, promising users a means of recovering crucial target RNA from samples of unprecedentedly low genomic quantity and quality.

And while this method was designed for RNA amplification related to HIV identification, Worobey believes this method will be of use not only for HIV and other viruses, but for a wide range of other applications such as detecting rare mutations that are known to contribute to certain cancers.

“We are right up against the limits of what has been possible,” he says. “We have broken new ground here; it will be useful not just for old samples and HIV-related studies, but for all sorts of clinical samples where HIV or other viruses or RNA targets may be at very low concentrations.”

To learn more about this technology, check out: Ultra-High Sensitivity RT-PCR for Viral Genome Sequencing (UA16-197)

To learn more about other RNA-related technologies from the University of Arizona, visit:

• Innovative Framework for Analyzing Genetic Data (UA15-065)
• A Device and Methods of the Use of Coated Particles for the Purification, Concentration, Management, and Storage of Biomolecules (UA05-015)

References:

[1] “What is RNA?” What is RNA?, RNA Society, www.rnasociety.org/about/what-is-rna/(link is external). Accessed 23 Feb. 2017.

[2] Ginsburg, Stephen. RNA amplification strategies for small sample populations. Elsevier, RNA amplification strategies for small sample populations, gene-quantification.com/ginsberg-singlecell-2005.pdf.

[3] “Dr. Michael Worobey, Department Head | Ecology and Evolutionary Biology.” Dr. Michael Worobey, Department Head | Ecology and Evolutionary Biology, University of Arizona Department of Ecology & Evolutionary Biology, eeb.arizona.edu/people/dr-michael-worobey-department-head. Accessed 22 Feb. 2017.

[4] Worobey, Michael, et al. “1970s and ‘Patient 0’ HIV-1 genomes illuminate early HIV/AIDS history in North America.” Nature, vol. 539, 3 Nov. 2016, pp. 98–101., doi:10.1038/nature19827. Accessed 23 Feb. 2017.