Generation of TLR7–/– mice. (A) Total bone marrow cell suspensions were prepared from WT or MyD88–/– mice and cultured for 18 h with media (open), 25 μg/ml poly(I:C) (cross-hatched), or 5 × 106 pfu/ml VSV (filled). IFNα levels were measured from culture supernatants by ELISA. (B) pDCs were purified from the bone marrow of WT mice by flow cytometric cell sorting and cultured with media (open) or 5 × 106 pfu/ml VSV (filled) for 18 h. IFNα levels were measured from culture supernatants by ELISA. (C) Structure of the TLR7 gene, the pZEN6 reporter cassette, and the targeted mutated locus. Black arrow heads denote loxP sites. (D) Northern blot analysis of RNA from WT and TLR7–/– bone marrow-derived macrophages. RNA was prepared from cells that were cultured for 4 h with media (–), 0.1 mg/ml lipopolysaccharide, or 100 nM R848. Ethidium bromide staining of the RNA gel is included as control (lower gel). (E) Whole-mount view of reporter gene expression (LacZ reporter, in blue) in mesenteric lymph nodes from TLR7–/– mice.