Table 1.
Details of the putative defense genes and primer sequences used for RT-PCR.
Fig 1.
a Phenotype of the lm3 mutant, the wildtype (3–1) plants and their F1 progeny at two weeks after anthesis in the field. b Comparison of lesion symptoms in different leaves in the lm3 mutant, the wildtype (3–1) plants and their F1 progeny. For each line, the top fourth, top third, top second and flag leaves are presented from left to right. c Lesion symptoms on the leaf sheath of the lm3 mutant, and wildtype (3–1). d-g Enhanced resistance to powdery mildew. d Reaction of the lm3 mutant and wildtype (3–1) plant to Blumeria graminis f. sp. tritici (Bgt) under natural infection in the adult plant stage under field conditions in 2009. The flag leaves of lm3 plants were covered with yellow necrotic spots, but no spores of Bgt were visible, while many white and brown powdery mildew spores were present on the wildtype (3–1) plants. e Powdery mildew reaction under natural infection at the adult plant stage in the greenhouse in 2015. Only two instances of Bgt sporulation were observed on the older leaves of lm3 plants (yellow arrows), but many white powdery mildew spores were visible on the wildtype (3–1) plants. f and g Responses of the lm3 mutant and wildtype (3–1) plants to Bgt E18 in the growth chamber at the adult stage in 2015. Many white and brown spores of Bgt were observed on the flag leaves (f, right) and top second leaves (g, right) leaves of wildtype (3–1) plants, whereas only one spore (yellow arrows) was present on each of the corresponding leaves of the lm3 mutant (f and g, left), where necrotic spots were widely distributed. All the visible brown spots on lm3 leaves and sheath are necrotic lesions, except a few spores of Bgt indicated by yellow arrows on lm3 leaves (e, f and g), while most visible whitish spots on the leaves of wildtype (3–1) plants are spores of Bgt on the panels (d, e, f and g).
Fig 2.
Chlorophyll content and ratio in the leaves of 3–1, lm3 and their F1 progeny.
a Chl a, Chl b and total Chl content in 3–1, lm3 and their F1 plants. b Ratio of Chl a to Chl b in 3–1, lm3 and their F1 plants. The means and SD (standard deviation) are shown with statistical analysis. Chl, chlorophyll; Chl a, chlorophyll a; Chl b, chlorophyll b; FW, fresh weight.
Fig 3.
Light dependence of lesion initiation.
a No lesion spots were observed in the middle segment of the leaves in the lm3 mutant, which was hidden from the light. b Effect of light intensity on lesion formation. Fewer lesion spots were observed under the treatment with 100 μmol m−2 s−1 (μE) compared with 200 μE in the lm3 mutant. c Effect of photoperiod on lesion formation. Several larger lesion spots were observed on the lm3 leaves after a longer day-length treatment. Two representative leaves are shown for each treatment, and the samples on the right are shown as the wildtype plant (3–1) in each panel.
Fig 4.
Histochemical analysis of wildtype and lesion-mimic plants.
a-d Trypan blue staining for cell death; e-h DAB staining for H2O2 accumulation. a, e Phenotype of the wildtype before Trypan blue and DAB staining; b, f Phenotype of the wildtype after Trypan blue and DAB staining; c, g Phenotype of the lesion mutant prior to Trypan blue and DAB staining, in which the brown spots are the lesions in the lm3 mutant; d, h Phenotype of the lesion mutant after Trypan blue and DAB staining, in which the dark blue (d) and reddish-brown spots (h) show the cell death and ROS enriched areas in the lm3 mutant, respectively. The data represent at least three experiments.
Fig 5.
Transcript abundance of defense-related genes during lesion formation.
Transcript abundance was determined via RT-qPCR, in samples of the roots (no lesions), the 1st leaves (fully developed lesions), and the 3rd leaves (developing lesions) of lm3 plants and corresponding tissues of 3–1 plants, and the fold-changes, corresponding to the relative expression of defense-related genes are shown on the y-axis, based on normalization of the expression data for lm3 to 3–1. The error bars represent the standard deviation between biological replicates. Mean fold-changes in the transcript abundance calculated using the △△Ct method between biological replicates ± standard deviation.
Table 2.
Segregation analysis of the LM symptoms in three segregating populations in 2007.
Fig 6.
Genetic map of the region surrounding lm3 on chromosome 3B.
a Map constructed using the DNAs from 190 individual F2 plants from a cross between lm3 and Jingdong8. b Detailed map of lm3 on chromosome 3B developed using the derived segregating population from the lm3/Jingdong8 cross, and the genome sequence of Chinese Spring. a and b are not in the same portrait.
Table 3.
Details of SSR markers used to map the lm3 locus.