Figure 2.

Induction of MTD in tobacco by treatment with INA or fungal elicitor. (A) MTD activity (Top), protein (Middle), and RNA (Bottom) in extracts from tobacco leaf discs, cultivar K326. Discs were floated on sterile distilled water either in the absence (−) or presence (+) of 1 mM INA for 48 hr before extraction. MTD activity was measured in extracts as above. Data represent mean ± SE of three independent experiments. Equal protein (20 μg) from each sample was separated by SDS/PAGE and blotted onto nitrocellulose. Blots were incubated with anti-MTD serum for 1 hr and visualized as described in Materials and Methods. Total RNA was also extracted from these tissue samples, and relative amounts of Mtd transcript were determined by blot analysis by using a ³²P-labeled, full-length celery Mtd cDNA (19) as a probe under conditions of moderate stringency (washed in 0.1× SSC at 50°C). (B) NT-1 tobacco cell suspensions were amended with either 1:50 (vol/vol) fungal elicitor (+; ref. 26) or an equal volume of sterile distilled water (−) and incubated for 12 hr. Cells were collected, and extracts were assayed for MTD activity (Upper) and protein (Lower) as described above. Data represent the means ± SE of three independent experiments.