Fig. 6.

Adv-CRISPR/AsCpf1 mediated CCR5 ablation suppresses HIV-1 infection in primary CD4⁺T cells. a The schematic diagram of the construction of CRISPR/AsCpf1 packaged adenovirus and corresponding transduction efficacy in primary CD4⁺T cells. b 300 ng of the CCR5 amplicons were identified with T7E1 after AdV-CRISPR/AsCpf1 transduction. c The CCR5 PCR products of the Adv-CRISPR/AsCpf1-#4/#5 transduced primary CD4⁺T cells were ligated with PGEM-T vector after poly A adding, and treated with DNA sequencing. d 10 μg of CCR5 protein of each group was assessed with western blotting. e R5-tropic HIV-1_YU-2 or X4-tropic HIV-1_NL4-3 was used to infect the adenovirus transduced primary CD4⁺T cells, the HIV-1 levels of each cells were determined from day 1 to day 7 post-infection. f Cell counts of each group were detected after the packaged adenovirus transduction for 7 days. g The apoptosis were analyzed with FCS at day 5 post-transduction. Necrotic cells (Annexin V-/7AAD +), necrotic or late apoptotic cells (Annexin V +/7AAD +); early apoptotic cells (Annexin V +/7AAD-); viable cells (Annexin V-/7AAD-). The data shown were the mean ± SD of three independent experiments. *P < 0.05; **P < 0.01; ***P < 0.001; NS, not significant; t test