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Acta Virologica Ahead of print Acta Virologica Vol.63, No.2, p.129-138, 2019
Acta Virologica Ahead of print Acta Virologica Vol.63, No.2, p.129-138, 2019
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Acta Virologica Vol.63, No.2, p.129-138, 2019 |
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| Title: A CRISPR/Cas9 library to map the HIV-1 provirus genetic fitness | ||
| Author: K. E. YODER | ||
| Abstract: The integrated proviral genome is the major barrier to a cure for HIV-1 infection. Genome editing technologies, such as CRISPR/Cas9, may disable or remove the HIV-1 provirus by introducing DNA double strand breaks at sequence specific sites in the viral genome. Host DNA repair by the error-prone non-homologous end joining pathway generates mutagenic insertions or deletions at the break. CRISPR/Cas9 editing has been shown to reduce replication competent viral genomes in cell culture, but only a minority of possible genome editing targets have been assayed. Currently there is no map of double strand break genetic fitness for HIV-1 to inform the choice of editing targets. However, CRISPR/Cas9 genome editing makes it possible to target double strand breaks along the length of the provirus to generate a double strand break genetic fitness map. We identified all possible HIV-1 targets with different bacterial species of CRISPR/Cas9. This library of guide RNAs was evaluated for GC content and potential off-target sites in the human genome. Complexity of the library was reduced by eliminating duplicate guide RNA targets in the HIV-1 long terminal repeats and targets in the env gene. Although the HIV-1 genome is AT-rich, the S. pyogenes CRISPR/Cas9 with the proto-spacer adjacent motif NGG offers the most HIV-1 guide RNAs. This library of HIV-1 guide RNAs may be used to generate a double strand break genetic fragility map to be further applied to any genome editing technology designed for the HIV-1 provirus. |
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| Keywords: HIV-1; genome editing; CRISPR; genetic fitness; guide RNAs | ||
| Year: 2019, Volume: 63, Issue: 2 | Page From: 129, Page To: 138 | |
| doi:10.4149/av_2019_201 |
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