Early Defense Mechanisms of Brassica oleracea in Response to Attack by Xanthomonas campestris pv. campestris

Black rot disease, caused by Xanthomonas campestris pv. campestris (Xcc), results in significant yield losses in Brassica oleracea crops worldwide. To find black rot disease-resistant cabbage lines, we carried out pathogenicity assays using the scissor-clipping method in 94 different B. oleracea lines. By comparing the lesion areas, we selected a relatively resistant line, Black rot Resistance 155 (BR155), and a highly susceptible line, SC31. We compared the two cabbage lines for the Xcc-induced expression pattern of 13 defense-related genes. Among them, the Xcc-induced expression level of PR1 and antioxidant-related genes (SOD, POD, APX, Trx H, and CHI) were more than two times higher in BR155 than SC31. Nitroblue tetrazolium (NBT) and diaminobenzidine tetrahydrochloride (DAB) staining analysis showed that BR155 accumulated less Xcc-induced reactive oxygen species (ROS) than did SC31. In addition, 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging assays showed that BR155 had higher antioxidant activity than SC31. This study, focused on the defense responses of cabbage during the early biotrophic stage of infection, indicated that Xcc-induced ROS might play a role in black rot disease development. We suggest that non-enzymatic antioxidants are important, particularly in the early defense mechanisms of cabbage against Xcc

Effect of cultivation factors on embryogenesis in isolated microspore culture of carrot (Daucus carota L.)

Using doubled haploid technologies inbreeding can significantly reduce the time to obtain homozygous parental lines required for the production of F1-hybrid of vegetable crops. This study aims to investigate the influence of factors on the efficiency of carrot embryogenesis in isolated microspore culture to optimise the elements of protocol for producing doubled haploids. Microspores were isolated from inflorescences of 21 genotypes and incubated in NLN13 medium supplemented with 0.1 mg·dm-3 2,4-dichlorophenoxyacetic acids, 0.1 mg·dm-3 1-naphthyl acetic acids, 130 g·dm-3 sucrose, and 400 mg·dm-3 casein hydrolysate and its modifications. Embryoids and their groups were formed after 2–6 months, in some cases after 12 months of cultivation. Depending on the variant, the embryogenesis efficiency averaged from 0 to 4.9 embryoids or groups of embryoids per Petri dish (10 cm3). Embryoids within the group were formed from different microspores. No significant effects of inflorescence position on the plant (branching order), sucrose, and casein hydrolysate concentration in the medium were observed. Significant advantages (p ≥ 0.05) for some genotypes were shown: 1) microspore suspension density 4·104 cells·cm-3 (5.0 embryoids per Petri dish were formed at a microspore suspension density of 4·104 cells·cm-3, 0.0 embryoids per Petri dish at a density of 8·104 cells·cm-3); 2) cultivating microspores of tetrad and early mononuclear stage (4.9 ±3.1 embryoids per Petri dish were obtained by culturing tetrads and early mononuclear microspores, while 0.6 ±0.7 embryoids per Petri dish were obtained by culturing of later developmental stages); 3) high-temperature treatment duration of five days (4.9 ±2.1 embryoids per Petri dish were obtained after five days of high-temperature treatment, 2.7 ±2.6 embryoids per Petri dish formed after two days of high-temperature treatment; 9.8 ±4.7, 10.1 ±6.1, 0.0 ±0.0 embryoids per Petri dish formed after two, five and eight days of high-temperature treatment respectively); 4) adding colchicine 0.5 mg·dm-3 to the nutrient medium for two days of high-temperature treatment, followed by medium replacement (3.3 ±2.6 embryoids per Petri dish were obtained by using a nutrient medium with colchicine, while 1.7 ±1.5 embryoids per Petri dish were obtained by culturing in the reference variant)