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Updated: Jun 5, 2026
Amplification of Near Full-length HIV-1 Proviruses for Next-Generation Sequencing
Published on: October 16, 2018
Multiple shRNA combinations for near-complete coverage of all HIV-1 strains
Glen J McIntyre1, Jennifer L Groneman, Yi-Hsin Yu
1Johnson and Johnson Research Pty Ltd, Level 4 Biomedical Building, 1 Central Avenue, Australian Technology Park, Eveleigh, NSW, 1430, Australia. glen@madebyglen.com.
Background:
Combinatorial RNA interference (co-RNAi) approaches are needed to account for viral variability in treating HIV-1 with RNAi, as single short hairpin RNAs (shRNA) are rapidly rendered ineffective by resistant strains. Current work suggests that 4 simultaneously expressed shRNAs may prevent the emergence of resistant strains.
Results:
In this study we assembled combinations of highly-conserved shRNAs to target as many HIV-1 strains as possible. We analyzed intersecting conservations of 10 shRNAs to find combinations with 4+ matching the maximum number of strains using 1220+ HIV-1 sequences from the Los Alamos National Laboratory (LANL). We built 26 combinations of 2 to 7 shRNAs with up to 87% coverage for all known strains and 100% coverage of clade B subtypes, and characterized their intrinsic suppressive activities in transient expression assays. We found that all combinations had high combined suppressive activities, though there were also large changes in the individual activities of the component shRNAs in our multiple expression cassette configurations.
Conclusion:
By considering the intersecting conservations of shRNA combinations we have shown that it is possible to assemble combinations of 6 and 7 highly active, highly conserved shRNAs such that there is always at least 4 shRNAs within each combination covering all currently known variants of entire HIV-1 subtypes. By extension, it may be possible to combine several combinations for complete global coverage of HIV-1 variants.
