Pseudovirus preparation and cell-entry assay
The pseudovirus cell-entry assay was performed as described previously
(Yang, Du et al. 2014). In brief, HEK293T cells were co-transfected with
a luciferase-expressing HIV-1 plasmid (pNL4-3.Luc.R-E-) and a plasmid
encoding HA-tagged SARS-CoV-BJ01 spike, SARS-CoV-2 spike or Pangolin CoV
spike. The supernatant containing pseudoviruses was collected 48 h after
transfection and the remaining cell pellet was lysed for Western blot
detection of HA-tagged spike proteins. In cell-entry assay,
pseudoviruses were incubated with recipient cells at 37 °C for 6 h, the
medium was changed and cells were incubated for an additional 42 h.
Cells were then washed with PBS buffer and lysed. Lysates were tested
for luciferase activity (Promega, USA). Each infection experiment was
carried out on for three times.