Integrated Proviral Sequencing Assay (IPSA)

University of Pittsburgh researchers have developed the Integrated Proviral Sequencing Assay (IPSA), a novel technology that allows for the rapid amplification and identification of rare viral integration sites within the human genome. This assay is particularly significant for studying retroviruses such as HIV-1 and HIV-2, as well as other viral DNA genomes like hepatitis B virus and human herpes virus type 6. IPSA’s ability to amplify and sequence entire viral genomes, along with flanking host sequences, represents a major advancement in understanding viral integration and its implications for human health.

Figure shows a schematic of the Integrated Proviral Sequencing Assay.

Description

The Integrated Proviral Sequencing Assay (IPSA) is designed to detect and sequence rare sites of viral integration in the human genome. Utilizing a novel DNA linker and optimized PCR techniques, IPSA achieves high specificity and sensitivity, overcoming the limitations of previous methods. The assay includes a partially double-stranded DNA oligo with a unique design to prevent non-specific amplification, ensuring accurate identification of viral integration sites. IPSA’s development involved extensive optimization to address challenges posed by secondary structures within viral genomes, enhancing the efficiency of enzymatic manipulations and PCR amplification.

Applications

• Research tool for genomics and virology
• HIV research and monitoring
• Viral integration site mapping
• Development of antiviral therapies

Advantages

IPSA offers high specificity and sensitivity in detecting rare viral integration sites, with the ability to sequence entire viral genomes along with flanking host sequences. It overcomes limitations of previous methods through optimized DNA linkers and PCR techniques, and is amenable to automation and scale-up using robotics and microfluidics. This technology provides critical insights into viral integration and persistence, aiding in the development of targeted antiviral therapies.

Invention Readiness

IPSA has undergone extensive development and optimization, demonstrating its effectiveness in detecting and sequencing viral integration sites. The method is ready for further validation and potential commercialization, with partial automation already achieved using robotic liquid handling and miniaturized sample processing.

IP Status

https://patents.google.com/patent/WO2020123787A1