RESUMEN
Clubroot disease caused by Plasmodiophora brassicae is becoming a serious threat to rapeseed (Brassica napus) production worldwide. Breeding resistant varieties using CR (clubroot resistance) loci is the most promising solution. Using marker-assisted selection and speed-breeding technologies, we generated Brassica napus materials in homozygous or heterozygous states using CRA3.7, CRA08.1, and CRA3.2 loci in the elite parental line of the Zhongshuang11 background. We developed three elite lines with two CR loci in different combinations and one line with three CR loci at the homozygous state. In our study, we used six different clubroot strains (Xinmin, Lincang, Yuxi, Chengdu, Chongqing, and Jixi) which are categorized into three groups based on our screening results. The newly pyramided lines with two or more CR loci displayed better disease resistance than the parental lines carrying single CR loci. There is an obvious gene dosage effect between CR loci and disease resistance levels. For example, pyramided lines with triple CR loci in the homozygous state showed superior resistance for all pathogens tested. Moreover, CR loci in the homozygous state are better on disease resistance than the heterozygous state. More importantly, no negative effect was observed on agronomic traits for the presence of multiple CR loci in the same background. Overall, these data suggest that the pyramiding of triple clubroot resistance loci conferred superior resistance with no negative effects on agronomic traits in Brassica napus.
Asunto(s)
Brassica napus , Resistencia a la Enfermedad , Enfermedades de las Plantas , Plasmodiophorida , Brassica napus/genética , Brassica napus/parasitología , Resistencia a la Enfermedad/genética , Enfermedades de las Plantas/parasitología , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/inmunología , Plasmodiophorida/fisiología , Plasmodiophorida/patogenicidad , Fitomejoramiento/métodos , FenotipoRESUMEN
The clubroot disease has become a worldwide threat for crucifer crop production, due to its soil-borne nature and difficulty to eradicate completely from contaminated field. In this study we used an elite resistant European fodder turnip ECD04 and investigated its resistance mechanism using transcriptome, sRNA-seq, degradome and gene editing. A total of 1751 DEGs were identified from three time points after infection, among which 7 hub genes including XTH23 for cell wall assembly and two CPK28 genes in PTI pathways. On microRNA, we identified 17 DEMs and predicted 15 miRNA-target pairs (DEM-DEG). We validated two pairs (miR395-APS4 and miR160-ARF) by degradome sequencing. We investigated the miR395-APS4 pair by CRISPR-Cas9 mediated gene editing, the result showed that knocking-out APS4 could lead to elevated clubroot resistance in B. napus. In summary, the data acquired on transcriptional response and microRNA as well as target genes provide future direction especially gene candidates for genetic improvement of clubroot resistance on Brassica species.
RESUMEN
Homogentisate Phytyltransferase (HPT) catalyzes condensation of homogentisate (HGA) and phytyl diphosphate (PDP) to produce tocopherols, but can also synthesize tocotrienols using geranylgeranyl diphosphate (GGDP) in plants engineered for deregulated HGA synthesis. In contrast to prior tocotrienol biofortification efforts, engineering enhanced tocopherol concentrations in green oilseeds has proven more challenging due to the integral role of chlorophyll metabolism in supplying the PDP substrate. This study show that RNAi suppression of CHLSYN coupled with HPT overexpression increases tocopherol concentrations by >two-fold in Arabidopsis seeds. We obtained additional increases in seed tocopherol concentrations by engineering increased HGA production via overexpression of bacterial TyrA that encodes chorismate mutase/prephenate dehydrogenase activities. In overexpression lines, seed tocopherol concentrations increased nearly three-fold, and resulted in modest tocotrienol accumulation. We further increased total tocochromanol concentrations by enhancing production of HGA and GGDP by overexpression of the gene for hydroxyphenylpyruvate dioxygenase (HPPD). This shifted metabolism towards increased amounts of tocotrienols relative to tocopherols, which was reflected in corresponding increases in ratios of GGDP/PDP in these seeds. Overall, our results provide a theoretical basis for genetic improvement of total tocopherol concentrations in green oilseeds (e.g., rapeseed, soybean) through strategies that include seed-suppression of CHLSYN coupled with increased HGA production.
RESUMEN
Oilseed rape (Brassica napus) has significant heterosis and Ogura CMS is a major way to use it. Ogura CMS has the advantages of complete and stable male sterility and easy-to-breed maintainers. Therefore, to breed better restorers has become an important goal for this system. Incidentally, clubroot is a soil-borne disease that is difficult to control by fungicidal chemicals, and it has been the main disease of oilseed rape in recent years in China, severely restricting the development of the oilseed rape industry. At present, the most effective method for controlling clubroot disease is to cultivate resistant varieties. One Ogura CMS restorer line (R2163) has shown much better combining ability, but lacks the clubroot disease resistance. This study was carried out to improve R2163 through marker-assisted backcross breeding (MABB). The resistant locus PbBa8.1 was introduced into the restorer R2163, and we then selected R2163R with clubroot disease resistance. Using the new restorer R2163R as the male parent and the sterile lines 116A and Z11A as the female parent, the improved, new resistant hybrids Kenyouza 741R and Huayouza 706R performed well, providing strong resistance and good agronomic traits. This work advances the utilization of heterosis and breeding for clubroot disease resistance in B. napus.
RESUMEN
As a vegetable oil, consisting principally of triacylglycerols, is the major storage form of photosynthetically-fixed carbon in oilseeds which are of significant agricultural and industrial value. Photosynthesis in chlorophyll-containing green seeds, along with photosynthesis in leaves and other green organs, generates ATP and reductant (NADPH and NADH) needed for seed fatty acid production. However, contribution of seed photosynthesis to fatty acid accumulation in seeds have not been well-defined. Here, we report the contribution of seed-photosynthesis to fatty acid production by probing segregating green (photosynthetically-competent) and non-green or yellow (photosynthetically-non-competent) seeds in siliques of an Arabidopsis chlorophyll synthase mutant. Using this mutant, we found that yellow seeds lacking photosynthetic capacity reached 80% of amounts of oil in green seeds at maturity. Combining this with studies using shaded siliques, we determined that seed-photosynthesis accounts for 20% and silique and leaf/stem photosynthesis each account for ~40% of the ATP and reductant for seed oil production. Transmission electron microscopy (TEM) and pyridine nucleotides and ATP analyses revealed that seed photosynthesis provides ATP and reductant for oil production mostly during early development, as evidenced by delayed oil accumulation in non-green seeds. Transcriptomic analyses suggests that the oxidative pentose phosphate pathway could be the source of carbon, energy and reductants required for fatty acid synthesis beyond the early stages of seed development.
RESUMEN
Clubroot is one of the most important diseases for many important cruciferous vegetables and oilseed crops worldwide. Different clubroot resistance (CR) loci have been identified from only limited species in Brassica, making it difficult to compare and utilize these loci. European fodder turnip ECD04 is considered one of the most valuable resources for CR breeding. To explore the genetic and evolutionary basis of CR in ECD04, we sequenced the genome of ECD04 using de novo assembly and identified 978 candidate R genes. Subsequently, the 28 published CR loci were physically mapped to 15 loci in the ECD04 genome, including 62 candidate CR genes. Among them, two CR genes, CRA3.7.1 and CRA8.2.4, were functionally validated. Phylogenetic analysis revealed that CRA3.7.1 and CRA8.2.4 originated from a common ancestor before the whole-genome triplication (WGT) event. In clubroot susceptible Brassica species, CR-gene homologues were affected by transposable element (TE) insertion, resulting in the loss of CR function. It can be concluded that the current functional CR genes in Brassica rapa and non-functional CR genes in other Brassica species were derived from a common ancestral gene before WGT. Finally, a hypothesis for CR gene evolution is proposed for further discussion.
Asunto(s)
Brassica napus , Brassica , Alimentación Animal , Brassica/genética , Brassica napus/genética , Mapeo Cromosómico , Genes prv , Filogenia , Fitomejoramiento , Enfermedades de las Plantas/genéticaRESUMEN
Clubroot caused by Plasmodiophora brassicae is a severe threat to the production of Brassica napus, worldwide. The cultivation of resistant varieties is the most efficient and environmentally friendly way to limit disease spread. We developed a highly resistant B. napus line, ZHE226, containing the resistance locus PbBa8.1. However, ZHE226 seeds contain high erucic acid content, which limits its cultivation owing to its low edible oil quality. A segregation population of BC3F2 was developed by crossing ECD04, a resistant European turnip donor, with Huangshuang5, an elite variety with no erucic acid in its seeds, as a recurrent plant. Fine mapping using the bulk segregation analysis sequencing (BSA-Seq) approach detected PbBa8.1 within a 2.9 MB region on chromosome A08. Interestingly, the previously reported resistance gene Crr1a was found in the same region. Genetic analysis revealed that the CAP-134 marker for Crr1a was closely linked with clubroot resistance (CR). Thus, PbBa8.1 and Crr1a might be allelic for CR. Moreover, comparative and genetic analysis showed that high erucic acid in the seeds of ZHE226 was due to linkage drag of fatty acid elongase 1 (FAE1) in the ECD04 line, which was located in the interval of PbBa8.1 with a physical and genetic distance of 729 Kb and 1.86 cm, respectively. Finally, a clubroot-resistant line with a low erucic acid content was successfully developed through gene-specific molecular marker assistant selection from BC4F4. These results will accelerate CR breeding programs in B. napus.