Linker-mediated PCR for global integration site analysis in splenic human CD4-positive T cells of HIV1-GFP-infected humanized mice

To comprehensively compare the HIV-1 integration sites between viral producing cells (GFP+ cells) and non-producing cells (GFP- cells), linker-mediated PCR is performed by using the samples obtained from HIV1-GFP-infected humanized mice. Additionally, to reactivate latently infected cells, viral non-producing cells (GFP- cells) were stimulated with PMA and ionomycin ex vivo. Then, the viral integration sites in potential latently infected cells (GFP-_GFP+ cells) and the cells with dead/inactivated proviruses (GFP-_GFP- cells) were analyzed by linker-mediated PCR. Overall design: GFP+ cells (n=13) and GFP- cells (n=13) were sorted from the splenic human CD4-positive T cells of HIV1-GFP-infected humanized mice at 2 weeks post-infection. GFP- cells were stimulated with PMA and ionomycin ex vivo, and then GFP+ cells (GFP-_GFP+ cells; n=6) and GFP- cells (GFP-_GFP- cells; n=6) were sorted. By using these samples, linker-mediated PCR was performed to describe genome-wide HIV-1 integration sites.