For every experiment, fresh plates were obtained by culturing the isolates from the 80 C stocks. M. perniciosa isolate ALF 42, obtained from green, infected branches at the municipality of Itajuı´pe, Bahia State, was used in the induction of resistance assays. Basidiospores of M. perniciosa were produced, collected, and stored according to Macagnan et al. (2005). Briefly, the fungus was cultivated on PDA for 10 days under laboratory conditions. Mycelium plugs were then transferred to petri dishes containing a sterile substrate composed of a homogeneous mixture of finely ground dry witches’ brooms (37%), oat flour (10%), CaSO4 (1.5%), and water (51.5%). Petri dishes were kept in an incubator at 25 C for 15 days. Fully colonized substrate was placed in a glass chamber (1.0 0.5 0.5 m) submitted to a watering regime of 1 l per day and a photoperiod of 8 h light and 16 h dark. After 15 days of incubation, the water supply was interrupted for 4 days to induce basidiocarp formation. The basidiocarps produced were collected, disinfested with streptomycin sulfate (150 lg per ml), washed with sterile distilled water, and had their caps fixed with Vaseline to the upper part of petri dishes. The lower part of the cap remained free for the discharge of basidiospores, which were collected in a solution containing glycerol (16%) and 2-(N-morpholine) ethane sulfonic (MES) acid (0.195%) with shaking for a period of 18 h under laboratory conditions. Basidiospore suspensions were kept in liquid nitro gen until use. Before use, the co ncentra tion of the basidio spore suspensi ons was adjusted to 5 105 basidiospores
per ml.
For every experiment, fresh plates were obtained by culturing the isolates from the 80 C stocks. M. perniciosa isolate ALF 42, obtained from green, infected branches at the municipality of Itajuı´pe, Bahia State, was used in the induction of resistance assays. Basidiospores of M. perniciosa were produced, collected, and stored according to Macagnan et al. (2005). Briefly, the fungus was cultivated on PDA for 10 days under laboratory conditions. Mycelium plugs were then transferred to petri dishes containing a sterile substrate composed of a homogeneous mixture of finely ground dry witches’ brooms (37%), oat flour (10%), CaSO4 (1.5%), and water (51.5%). Petri dishes were kept in an incubator at 25 C for 15 days. Fully colonized substrate was placed in a glass chamber (1.0 0.5 0.5 m) submitted to a watering regime of 1 l per day and a photoperiod of 8 h light and 16 h dark. After 15 days of incubation, the water supply was interrupted for 4 days to induce basidiocarp formation. The basidiocarps produced were collected, disinfested with streptomycin sulfate (150 lg per ml), washed with sterile distilled water, and had their caps fixed with Vaseline to the upper part of petri dishes. The lower part of the cap remained free for the discharge of basidiospores, which were collected in a solution containing glycerol (16%) and 2-(N-morpholine) ethane sulfonic (MES) acid (0.195%) with shaking for a period of 18 h under laboratory conditions. Basidiospore suspensions were kept in liquid nitro gen until use. Before use, the co ncentra tion of the basidio spore suspensi ons was adjusted to 5 105 basidiosporesper ml.
การแปล กรุณารอสักครู่..