Clubroot is a devastating disease of majority of the members Brassicaceceae family. Chinese cabbage is a member of Brassica rapa family that become severely affected with clubroot disease in Korean fields. However, molecular investigations towards detection of the causal agent of this disease, Plasmodiophora brassicae, are completely missing in Korea. Possible reasons are the complicated life cycle of the pathogen and genome similarity with its host pathogen. With a view to detect the field isolates were collected from nine different locations of Korea, in the first experiment ribosomal DNA (rDNA) sequence of 11 isolates were cloned and sequenced. Afterwards, ten HRM (high resolution melting) probes were designed based on variability in rDNA sequences exists in field isolates. Experiment 1, revealed that there were major nucleotide sequence variations in intron1 of the smaller subunit of rDNA of 11 field isolates. A 280 bp deleted sequences were resulted in some isolates along with original 657 bp sequences. Four HRM probes, out of ten probes, were able to detect and separate Korean P. brassicae isolates into two groups for notable melting temperature variation. Among the eleven Korean field isolates; ‘Seosan’ isolate was a pathotype 4 group isolate that was found virulent against Japanese clubroot resistant (CR) cultivar bearing Crr1, Crr2 and CRb resistance genes. This study, therefore, hypothesized that a resistant genotype (turnip) against ‘Seosan’ isolate could be a source of a novel resistant gene against this pathogen. In the second experiment it was planned to identify and map the resistant QTL (quantitative trait loci) in a segregating population between CR turnip and susceptible Chinese cabbage using RAD-seq technology. The resistant QTL xii
was eventually mapped that bears 11 candidate genes with disease resistance related, NBS-LRR or protein kinase domains. A total of 17 HRM probes were tested in this study to develop suitable markers that can effectively screen Seosan-isolate resistant Chinese cabbage genotypes. Four HRM markers clearly separated resistant and susceptible segregating genotypes in a similar fashion. In the third experiment, this study explored the possible role of cytokinin biosynthesis genes in clubroot formation. Expression of 17 cytokinin biosynthesis genes was investigated in leaf and root samples of both inoculated and non-inoculated plants at five different time-points (1, 3, 14, 28 and 35 days) after inoculation. Number of genes in root tissues during gall expansion at 35 days after was very less compared to leaf tissues indicating that during gall expansion the pathogen induced cytokinin-like substances became activated. The fourth experiment was designed to understand genetic and biochemical underpinning of wax formation in cabbage lines. This study explored the expression pattern of wax biosynthesis genes in high-wax and low-wax cabbage lines at their three different growth phases and also investigated the ultra-morphology and biochemical composition of wax crystals. This study revealed that wax biosynthesis genes become activated at the very early stage of cabbage plants. The relative transcript abundance was the highest at the leaf position four (about 20 days old) and that greatly declined at the older, senescing leaves. Expression of wax biosynthesis genes, especially that of BoCER3 gene, was higher in high-wax lines compared to low-wax lines and that probably resulted in prominent and large wax crystals in high-wax lines compared to low-wax lines. C29-alkanes, ketones and secondary alcohols were predominant in both high- and low-wax lines but high-wax lines measured significantly higher amount of total waxes per unit leaf area. The new knowledge generated in this study will be helpful in developing clubroot resistant Brassica crops and also aid effort developing cabbage lines with desired levels of leaf waxes that would be able to tackle a number of biotic and abiotic stresses.