title: Silent and expressed HIV-1 in resting and active CD4+ T-cells
design: Pace et al. (2012) spinoculated CD4+ T cells with HIV NL4-3 at an MOI of 0.1. After 96 hours, the cells were stained for intracellular Gag CD25, CD69 and HLA-DR and sorted into four subpopulations based on activation state and Gag expression; activated Gag-, activated Gag+, resting Gag- and resting Gag+. The ability of the viruses to reactivate was not tested although previous studies have shown that the majority are likely inducible (Plesa et al., 2007). Genomic DNA was extracted and digested with restriction enzymes MseI and Tsp509 and ligated to adapters. Proviral LTR-host genome junctions were sequenced by 454 pyrosequencing after nested PCR.