|
|
Mapping of clubroot resistance genes by associative transcriptomics
HEJNA, Ondřej
The current high pathogenic pressure on agriculture crops is the task for breeders. One of the serious problems that modern breeding is struggling to address is clubroot. This disease is caused by the soil biotrophic pathogen Plasmodiophora brassicae. At present, this pathogen presents serious problems for most cruciferous crops worldwide. The possibilities for controlling this disease are very limited. From an economic point of view, the most effective option is growing resistant or tolerant varieties against this pathogen. To date, several genetic sources of resistance for breeding have been identified. However, the application of these resources inbreeding is problematic, because there is a lack of knowledge of the basis of the resistance and lack of understanding of causal genes or proteins. In this study, the resistance of the B. napus genotypes to clubroot was evaluated. An Associative Transcriptomics approach was used to identify markers and causal genes associated with resistance to clubroot. In assessing the response of a genetic diversity panel comprising 250 accessions of B. napus, the mixed pathotype ECD 17/31/31 was used. The result of this testing finds 46 genotypes with disease index (DI) greater than 80. Such varieties can be considered resistant. For 245 genotypes, RNAseq transcriptome data were obtained for use in two associative analyses. In the first assay, SNP association analysis was performed with 256,397 SNP markers distributed across the genome of B. napus. The result of this analysis revealed 86 SNP markers located in 9 different regions on chromosomes A01, A02, A03, A08, C02 and C07. Resistant loci were named according to their location as BnA01_0308, BnA02_0265, BnA02_0286, BnA03_0186, BnA03_0263, BnA08_0009, BnCO02_0414, BnCO07_0238 and BnCO07_0421. Altogether, 392 genes were found in these regions. For the second association analysis, transcript abundance data of 53,889 genes in the B. napus genome were used. The analysis itself evaluated the degree of association between expression profiles and DI profile. Based on this analysis, 19 associated genes were revealed. In the last step, annotation analyzes GO, InterPro, and annotations were performed based on similarity to Arabidopsis thaliana genes. Overall, 82 candidate genes were selected to be involved in clubroot resistance. These results provide useful information both for breeding by identifying associated SNP markers useful for breeding resistant varieties and for understanding the elements of resistance in B. napus.
|
|
|
The characterisation of organ-specific phytohormone responses to nutrient deficiency and biotic stress
Kramná, Barbara ; Vaňková, Radomíra (advisor) ; Hronková, Marie (referee) ; Plíhalová, Lucie (referee)
Abiotic and biotic stresses lead to crop yield losses and ultimately negatively affect agriculture production. Elucidation of the mechanisms of plant stress responses and their regulation could help to understand plant defence and improve stress tolerance. Phytohormones stand behind both plant growth and developmental changes as well as stress signalling. This thesis summarises the results published in two articles focused on phytohormone dynamics in response to abiotic and biotic stresses, namely phosphate shortage and Plasmodiophora brassicae infection. A review article focuses in depth on strigolactones as master regulators of phosphate deficiency responses. The main emphasis is put on organ-specific reactions and exogenous phytohormone treatment with the potential to convey stress tolerance. In the case of phosphate shortage, the universal reaction in all organs was a decrease in active cytokinin trans-zeatin and gibberellin GA4 with a concurrent elevation of abscisic acid. Also, the high- affinity phosphate transporters (PHT1;4 and PHT1;7) exhibited increased gene expression within the whole plant. Shoot apical meristems showed numerous changes in gene expression and were the most affected organ during the lack of phosphate. Only in roots, we observed a substantial elevation of low active...
|
|
|
The characterisation of organ-specific phytohormone responses to nutrient deficiency and biotic stress
Kramná, Barbara ; Vaňková, Radomíra (advisor) ; Hronková, Marie (referee) ; Plíhalová, Lucie (referee)
Abiotic and biotic stresses lead to crop yield losses and ultimately negatively affect agriculture production. Elucidation of the mechanisms of plant stress responses and their regulation could help to understand plant defence and improve stress tolerance. Phytohormones stand behind both plant growth and developmental changes as well as stress signalling. This thesis summarises the results published in two articles focused on phytohormone dynamics in response to abiotic and biotic stresses, namely phosphate shortage and Plasmodiophora brassicae infection. A review article focuses in depth on strigolactones as master regulators of phosphate deficiency responses. The main emphasis is put on organ-specific reactions and exogenous phytohormone treatment with the potential to convey stress tolerance. In the case of phosphate shortage, the universal reaction in all organs was a decrease in active cytokinin trans-zeatin and gibberellin GA4 with a concurrent elevation of abscisic acid. Also, the high- affinity phosphate transporters (PHT1;4 and PHT1;7) exhibited increased gene expression within the whole plant. Shoot apical meristems showed numerous changes in gene expression and were the most affected organ during the lack of phosphate. Only in roots, we observed a substantial elevation of low active...
|
|
|
Resistance of cole vegetable cultivars against Plasmodiophora brassicae pathotypes in Czech Republic
Zacharová, Lea ; Kazda, Jan (advisor) ; Gerhard, Gerhard (referee)
Clubroot disease of crucifer plants is caused by a pathogen Plasmodiophora brassicae (Wor.). It is counted among the most common and most serious diseases that attack not only rape, but also, for example, head cabbage, cauliflower, broccoli and other kinds of cabbage vegetables.
Plasmodiophora brassicae is descirebed as a very durable and well-developed body in this thesis. Physical, chemical and biological components of the soil environment are in the thesis associated with an effect on survival, growth and reproduction of this microbe. Obviously, Plasmodiophora's brassicae successfulness is given due to its resting spores and because of them the microbe is able to survive in a soil for years.
The experimental part of my thesis is focused on a testing of resistance of selected cultivars against Plasmodiophora brassicae. Each variety were grown under the control within conditions in the greenhouse. To evaluate the degree of a damage of the plants scale of Buzacki (1975), Strelkov et al. (2007) were applied, the index of an invasion was calculated based on these scales.
Cultivars were exposed to a pathotype 6 and as a control variety was used the cultivar Granaat. The results had shown the most susceptible respond of a cultivar red cabbage F1 Loder and other varieties tested had confirmed their resistance to Plasmodiophora brassicae.
|
|
|
Plasmodiophora brassicae on winter rape
Řičařová, Veronika ; Ryšánek, Pavel (advisor) ; Jaroslav, Jaroslav (referee)
Winter oilseed rape (Brassica napus) is an important crop in the Czech Republic. Clubroot disease caused by the pathogen Plasmodiophora brassicae Wor. is a serious and still-growing problem for oilseed rape growers. Research on P. brassicae in the Czech Republic is therefore important for the development of effective strategies to manage clubroot under Czech environmental conditions.
One of the aims of this study was monitoring of this pathogen. The disease was previously widespread in commercial vegetable production and in hobby gardens. Since 2010, oilseed rape clubroot started to spread across the whole country, whereas it had previously only been observed in the northeast. Clubroot occurrence was monitored for five years by the Union of Oilseed Growers and Processors on the basis of disease symptoms present on oilseed rape fields. The presence of P. brassicae and clubroot symptoms were reported in all regions of the Czech Republic, except the Ústecký Region, and in 31 out of 76 districts. At present, at least 130 fields are known to be infested by the pathogen, but this number is very likely underestimated. Some soil samples were also tested by conventional PCR (polymerase chain reaction) to evaluate the possibility of their usage. All 14 suspected samples tested positive by PCR.
The next aim was to evaluate the pathotype composition of P. brassicae populations from the Czech Republic, according to the three evaluation systems, and to determine soil inoculum loads for representative fields via traditional end-point PCR as well as quantitative PCR analysis. There were considerable differences between the populations of P. brassicae, and the number of pathotypes varied depending on the evaluation system and the threshold used to distinguish susceptible vs. resistant plant reactions. This is the first study comparing the effect of different thresholds. Using an index of disease (ID) of 25 % to distinguish susceptible vs. resistants reactions, there was a total of five pathotypes identified based on the differentials of Williams, five with the system of Somé et al., and 10 with the European Clubroot Differential (ECD) set. However, based on a threshold of 50%, there were five pathotypes according to the evaluation system by Williams, four based on the differentials of Somé et al. and 8 with the ECD set. Changing of the thresholds led to the reclassification of some pathotypes. Pathotypes 7 by Williams was the most frequent in both thresholds. High amounts of pathogen DNA were found in many of the field soils analysed by quantitative PCR.
Experiments with P. brassicae-resistant cultivars of winter oilseed rape were conducted in an infested field and greenhouse. In the greenhouse, six resistant cultivars were grown in infested soil collected from various fields in the Czech Republic and assessed for index of disease (ID %). The best results bring cultivar Mentor (2+- 0.7 %) closely followed by cultivar SY Alister (5+-1.1 %), the highest ID had cultivar CHW 241 (30+-3.8%). In the field experiment, seven resistant cultivars were grown, and disease development was monitored monthly. The lowest index of disease brought cultivar Andromeda (3+- 0.8 %) and PT 235 (4+-1.5 %), the highest ID has cultivar CWH 241(46 +- 6.5 %) in the first season and in the second season any cultivar achieved 25 % ID. Yields were measured at the end of the cropping season. The highest yield was achieved by cultivar SY Alister (6.1 t/ha) in the first season and cultivar PT 242 (5.03 t/ha) in the second season. The inoculum level was measured across the field by (qPCR), and a map of the infestation was created. The highest spore concentration was found on the field entrance. Collectively, the information obtained on the effectiveness of host resistance and pathogenic diversity of P. brassicae populations from the Czech Republic may help to more effectively manage clubroot in this country.
|
guest ::
