FIG 8.

Wheat streak mosaic virus (WSMV) coat protein (CP) amino acids 101 to 300 induce superinfection exclusion. (A and B) Superinfection of WSMV-GFP on wheat systemically infected with TriMV expressing WSMV CP or CP deletions. TriMV and WSMV, wheat systemically infected with wild-type TriMV and WSMV, respectively. Buffer and buffer-NC, buffer-inoculated wheat subsequently challenge inoculated or not challenge inoculated with WSMV-GFP, respectively. Images shown are local foci at 5 days post-challenge inoculation (dpci) (A) and systemic infection at 9 dpci (B) by WSMV-GFP (challenge virus). Bars, 500 μm. (C and D) Western blot assay of total proteins from the challenge virus-inoculated leaves at 5 dpci (C) and upper noninoculated leaves at 9 dpci (D). Full-length and truncated TriMV CP are indicated with arrows and arrowheads, respectively. Asterisks indicate a wheat protein reacting nonspecifically with TriMV polyclonal antiserum. Presented Coomassie blue-stained SDS-PAGE gels show the large subunit of wheat RubisCO protein for the amount of protein loaded per well in Western blot assays. (E and F) Ethidium bromide-stained agarose gels of RT-PCR products with TriMV-specific primers on either side of insert showing the stability of inserted sequences at 5 dpci of inoculated (E) and 9 dpci of upper noninoculated (F) leaves. (C to F) Lanes 1 to 8, wheat infected with TriMV (lane 1) or TriMV expressing WSMV full-length CP (lane 2) or CP with deletion of amino acids 3 to 35 (lane 3), 36 to 84 (lane 4), 85 to 100 (lane 5), 101 to 200 (lane 6), 201 to 300 (lane 7), and 294 to 349 (lane 8); lane 9, TriMV expressing WSMV CP amino acids 101 to 300; lane 10, wheat infected with WSMV; lane 11, buffer-inoculated wheat subsequently challenge inoculated; lane 12, buffer-inoculated wheat with no challenge inoculation; lane +, pTriMV as a positive control; lanes M, 1.0-kbp DNA ladder.