Wenxing Pang

Genome-wide Analysis and Characterization of Aux/IAA Family Genes in Brassica rapa

Auxins are the key players in plant growth development involving leaf formation, phototropism, root, fruit and embryo development. Auxin/Indole-3-Acetic Acid (Aux/IAA) are early auxin response genes noted as transcriptional repressors in plant auxin signaling. However, many studies focus on Aux/ARF gene families and much less is known about the Aux/IAA gene family in Brassica rapa (B. rapa). Here we performed a comprehensive genome-wide analysis and identified 55 Aux/IAA genes in B. rapa using four conserved motifs of Aux/IAA family (PF02309). Chromosomal mapping of the B. rapa Aux/IAA (BrIAA) genes facilitated understanding cluster rearrangement of the crucifer building blocks in the genome. Phylogenetic analysis of BrIAA with Arabidopsis thaliana, Oryza sativa and Zea mays identified 51 sister pairs including 15 same species (BrIAA—BrIAA) and 36 cross species (BrIAA—AtIAA) IAA genes. Among the 55 BrIAA genes, expression of 43 and 45 genes were verified using Genebank B. rapa ESTs and in home developed microarray data from mature leaves of Chiifu and RcBr lines. Despite their huge morphological difference, tissue specific expression analysis of BrIAA genes between the parental lines Chiifu and RcBr showed that the genes followed a similar pattern of expression during leaf development and a different pattern during bud, flower and siliqua development stages. The response of the BrIAA genes to abiotic and auxin stress at different time intervals revealed their involvement in stress response. Single Nucleotide Polymorphisms between IAA genes of reference genome Chiifu and RcBr were focused and identified. Our study examines the scope of conservation and divergence of Aux/IAA genes and their structures in B. rapa. Analyzing the expression and structural variation between two parental lines will significantly contribute to functional genomics of Brassica crops and we belive our study would provide a foundation in understanding the Aux/IAA genes in B. rapa.

Genetic Detection of Clubroot Resistance Loci in a New Population of Brassica rapa

Clubroot is one of the most serious diseases affecting Brassica crop production worldwide. In order to identify the location of clubroot resistance genes in Chinese cabbage, we constructed a linkage map for an F2 population derived from a cross between a resistant turnip inbred line, ‘Siloga’ (B. rapa ssp. rapifera), and a susceptible Chinese cabbage inbred line, ‘BJN3’ (B. rapa ssp. pekinensis). The newly developed genetic map included 207 markers and covered 1151.7 cM. In combination with clubroot resistance tests from the field, the data allowed us to identify three quantitative trait loci of clubroot resistance, using a composite interval mapping method. Clubroot resistance genes QS_B1.1, QS_B3.1, and QS_B8.1 were located on chromosomes A1, A3, and A8, respectively, of B. rapa. Their contribution rates were 8.18%, 70.55%, and 7.28%, respectively. QS_B1.1 was a novel locus of clubroot resistance, independent of any published clubroot-resistance loci. QS_B3.1 was first detected in ‘Siloga,’ mapped to the previous CRb and CRa region, whereas QS_B8.1 was closely linked to Crr1b. Epistatic interactions with additive effects were detected between QS_B3.1 and QS_B8.1.

Quantitative Trait Loci for Morphological Traits and their Association with Functional Genes in Raphanus sativus

Identification of quantitative trait loci (QTLs) governing morphologically important traits enables to comprehend their potential genetic mechanisms in the genetic breeding program. In this study, we used 210 F2 populations derived from a cross between two radish inbred lines (Raphanus sativus) “835” and “B2,” including 258 SSR markers were used to detect QTLs for 11 morphological traits that related to whole plant, leaf, and root yield in 3 years of replicated field test. Total 55 QTLs were detected which were distributed on each linkage group of the Raphanus genome. Individual QTLs accounted for 2.69–12.6 of the LOD value, and 0.82–16.25% of phenotypic variation. Several genomic regions have multiple traits that clustered together, suggested the existence of pleiotropy linkage. Synteny analysis of the QTL regions with A. thaliana genome selected orthologous genes in radish. InDels and SNPs in the parental lines were detected in those regions by Illumina genome sequence. Five identified candidate gene-based markers were validated by co-mapping with underlying QTLs affecting different traits. Semi-quantitative reverse transcriptase PCR analysis showed the different expression levels of these five genes in parental lines. In addition, comparative QTL analysis with B. rapa revealed six common QTL regions and four key major evolutionarily conserved crucifer blocks (J, U, R, and W) harboring QTL for morphological traits. The QTL positions identified in this study will provide a valuable resource for identifying more functional genes when whole radish genome sequence is released. Candidate genes identified in this study that co-localized in QTL regions are expected to facilitate in radish breeding programs.

Development of a leafy Brassica rapa fixed line collection for genetic diversity and population structure analysis

Brassica rapa is an economically important crop with a wide range of morphologies. Developing a set of fixed lines and understanding their diversity has been challenging, but facilitates resource conservation. We investigated the genetic diversity and population structure of 238 fixed lines of leafy B. rapa with 45 new simple sequence repeat markers and 109 new NGS (next-generation sequencing)-generated single nucleotide polymorphism markers evenly distributed throughout the B. rapa genome. Phylogenetic analysis classified the vegetable fixed lines into four subgroups, with the three oil types forming a separate and relatively distant cluster. A model-based population structure analysis identified four subpopulations corresponding to geographical origins and morphological traits, and revealed extensive allelic admixture. In particular, the Chinese cabbage cluster was subdivided into three groups and showed considerable correlation with leaf- and heading-related traits (leaf and heading shape). The vegetable B. rapa fixed lines successfully developed in our study could be valuable materials for establishing a multinational Brassica rapa diversity resource. Understanding the genetic diversity and population structure could be useful for utilization of the representative genetic variation and further genomic analysis.

Anatomic Characteristics Associated with Head Splitting in Cabbage (Brassica oleracea var. capitata L.).

Cabbage belonging to Brassicaceae family is one of the most important vegetables cultivated worldwide. The economically important part of cabbage crop is head, formed by leaves which may be of splitting and non-splitting types. Cabbage varieties showing head splitting causes huge loss to the farmers and therefore finding the molecular and structural basis of splitting types would be helpful to breeders. To determine which anatomical characteristics were related to head-splitting in cabbage, we analyzed two contrasting cabbage lines and their offspring using a field emission scanning electron microscope. The inbred line “747” is an early head-splitting type, while the inbred line “748” is a head-splitting-resistant type. The petiole cells of “747” seems to be larger than those of “748” at maturity; however, there was no significant difference in petiole cell size at both pre-heading and maturity stages. The lower epidermis cells of “747” were larger than those of “748” at the pre-heading and maturity stages. “747” had thinner epidermis cell wall than “748” at maturity stage, however, there was no difference of the epidermis cell wall thickness in the two lines at the pre-heading stage. The head-splitting plants in the F1 and F2 population inherited the larger cell size and thinner cell walls of epidermis cells in the petiole. In the petiole cell walls of “747” and the F1 and F2 plants that formed splitting heads, the cellulose microfibrils were loose and had separated from each other. These findings verified that anomalous cellulose microfibrils, larger cell size and thinner-walled epidermis cells are important genetic factors that make cabbage heads prone to splitting.

The effects of fluctuating culture temperature on stress tolerance and antioxidase expression in Esteya vermicola

The endoparasitic nematophagous fungus, Esteya vermicola, has shown great potential as a biological control agent against the pine wood nematode, Bursaphelenchus xylophilus. Fluctuating culture temperatures can affect fungal yields and fungal tolerance to desiccation, UV radiation, H2O2, and heat stress, as well as antioxidase expression. To explore these effects, E. vermicola cultured under five temperature ranges, 26°C, 15–26°C, 26–35°C, 20–30°C, and 15–35°C, were compared. The cultures grown at lower temperatures showed better growth, stronger tolerance to desiccation, UV, and H2O2 stresses, and increased catalase expression, However, these cultures also showed weaker heat stress tolerance and lower superoxide dismutase expression than the higher-temperature cultures. In particular, the E. vermicola cultured at 20–30°C, i.e., fluctuating in a narrow range around the optimal temperature, showed the best performance. Therefore, for production in practical applications, this narrowly fluctuating, moderate temperature appears to be optimal for yield and stress tolerance in E. vermicola.

Mining of Brassica-Specific Genes (BSGs) and Their Induction in Different Developmental Stages and under Plasmodiophora brassicae Stress in Brassica rapa

Orphan genes, also called lineage-specific genes (LSGs), are important for responses to biotic and abiotic stresses, and are associated with lineage-specific structures and biological functions. To date, there have been no studies investigating gene number, gene features, or gene expression patterns of orphan genes in Brassica rapa. In this study, 1540 Brassica-specific genes (BSGs) and 1824 Cruciferae-specific genes (CSGs) were identified based on the genome of Brassica rapa. The genic features analysis indicated that BSGs and CSGs possessed a lower percentage of multi-exon genes, higher GC content, and shorter gene length than evolutionary-conserved genes (ECGs). In addition, five types of BSGs were obtained and 145 out of 529 real A subgenome-specific BSGs were verified by PCR in 51 species. In silico and semi-qPCR, gene expression analysis of BSGs suggested that BSGs are expressed in various tissue and can be induced by Plasmodiophora brassicae. Moreover, an A/C subgenome-specific BSG, BSGs1, was specifically expressed during the heading stage, indicating that the gene might be associated with leafy head formation. Our results provide valuable biological information for studying the molecular function of BSGs for Brassica-specific phenotypes and biotic stress in B. rapa.

The 2015 KSM-ICWG-GSP Joint Clubroot Symposium Meeting Report

Clubroot disease is one of the most damaging diseases worldwide for Brassica crops. Researchers come together during various meetings to report on their findings to understand or combat the disease. In 2015, the Korean Society of Mycology together with the International Clubroot Working Group and the Golden Seed Project were hosts for the clubroot symposium in Daejeon, Korea. Presentations during the conference ranged from basic research, for example, the genome sequence of the clubroot pathogen Plasmodiophora brassicae, but also transcriptome and metabolome studies. Further studies on the biology of P. brassicae are essential to understand the nature of disease development and interaction with plants. Major topics discussed throughout the symposium were the generation of single-spore isolates and breeding of resistant cultivars. In addition, it was reported that biocontrol agents are worth investigating for their capability to control clubroot.