High hydrostatic pressure has been demonstrated very useful in many of the steps of nucleic acid sample preparation including cell lysis, deproteination, nuclease inhibition and nucleic acid extraction from various biological samples. During this Phase I study, feasibility of extraction of RNA from plasma will be examined using BioSeq's innovative high-pressure technology. HIV 8E5, a non-infectious HIV mutant virus, will be employed as the target RNA source. Profiles of pressure, temperature, and addition of necessary chemicals will be tested to select the optimum conditions for viral lysis, nuclease inactivation and RNA extraction. Qualitative and quantitative nucleic acid assays will be performed for each step and compared to those from the methods that are commonly used. The pressure extraction system as well as the conceptual design of the extraction module and the possibility of integration will be developed. Data generated in this phase of study will form the basis for the design of a rapid, automated system for RNA and DNA isolation during the Phase II. The proposed pressure-assisted RNA extraction would reduce the usage of harsh chemical reagents, labor and material cost, and assay time. It represents an important advancement in the molecular diagnostics industry for monitoring infectious diseases.
Nucleic acid extraction using high-pressure cycling is an innovative process. Viral RNA extraction from plasma proposed here aims to overcome some of the current limitations of sample preparation in molecular diagnostics. The development of this pressure technology will ultimately permit rapid, automated nucleic acid sample preparation without the requirement of extensive dilution of the sample or use of harsh chemicals. The success of the proposed technology will benefit not only viral diagnostics, but also broader fields, such as infectious disease diagnostics, cancer diagnosis, genomics and forensic analysis for research and clinical applications.
