Jingbin Jiang

Mapping and candidate gene screening of tomato Cladosporium fulvum-resistant gene Cf-19, based on high-throughput sequencing technology.

BackgroundTomato leaf mold is a common disease in tomato cultivation. This disease is caused by Cladosporium fulvum, which has many physiological races and differentiates rapidly. Cf genes confer resistance to C. fulvum, and the C. fulvum-tomato pathosystem is a model for the study of gene-for-gene interactions. Plants carrying the Cf-19 gene show effective resistance to C. fulvum in the field, and can be used in breeding and resistance mechanism studies as new resistant materials. In this study, we used F2 bulk specific-locus amplified fragment sequencing (SLAF-seq) and parental resequencing methods to locate and characterize the Cf-19 gene.ResultsA total of 4108 Diff_markers and three association regions were found in association analysis. A 2.14-Mb region containing seven Cf-type genes was identified in further analysis based on data from SLAF-seq and parental resequencing. Two candidate genes, Solyc01g006550.2.1 and Solyc01g005870.1.1, were screened out by quantitative real-time PCR (qRT-PCR) analysis. Sequence analysis showed that Solyc01g006550.2.1 (an allelic locus of Cf-0) in CGN18423 was a novel homologue of the Cladosporium resistance gene Cf-9 (Hcr9s) in the Cf-4/9 locus. The marker P7, which cosegregated with the resistant trait, was developed based on sequence mutation of the Solyc01g006550.2.1 locus in CGN18423.ConclusionsThe Cf-19 gene was mapped to the short arm of chromosome 1. The candidate genes Solyc01g006550.2.1 and Solyc01g005870.1.1 showed related amino acid sequence structures and expression patterns. Solyc01g006550.2.1 had a close evolutionary relationship with the functional Hcr9 members Cf-4 and Cf-9, and was very different from non-functional members. The results from this study will facilitate the breeding of cultivars carrying the Cf-19 gene and provide a basis for further gene cloning, resistance gene evolution and plant resistance mechanism studies.

Transcriptome Analysis of the Sm-Mediated Hypersensitive Response to Stemphylium lycopersici in Tomato

Gray leaf spot disease caused by Stemphylium lycopersici is a major disease in cultivated tomato plants and threatens tomato-growing areas worldwide. Sm is a single dominant gene that confers resistance to tomato gray leaf spot disease agent. However, the underlying molecular mechanism remains unclear. Here, resistant (cv. Motelle, containing the Sm gene) and susceptible (cv. Moneymaker) plants were inoculated with virulent Stemphylium lycopersici isolate at a time point at which both cultivars showed a strong response to S. lycopersici infection. Transcriptome analyses were performed in both cultivars using RNA-seq. The number of differentially expressed genes (DEGs) was higher in Motelle than Moneymaker. Functional classification revealed that most DEGs were involved in plant–pathogen interactions, plant hormone signal transduction, regulation of autophagy, glycerophospholipid metabolism, and α-linolenic acid metabolism. Moreover, the genes that were significantly up-regulated in Sm tomatoes were involved in plant–pathogen interaction pathways. A total of 26 genes were selected for confirmation of differentially expressed levels by quantitative real-time PCR. This knowledge will yield new insights into the molecular mechanism of Sm responses to S. lycopersici infection.

Transcriptome Analysis of the Cf-12-Mediated Resistance Response to Cladosporium fulvum in Tomato

Cf-12 is an effective gene for resisting tomato leaf mold disease caused by Cladosporium fulvum (C. fulvum). Unlike many other Cf genes such as Cf-2, Cf-4, Cf-5, and Cf-9, no physiological races of C. fulvum that are virulent to Cf-12 carrying plant lines have been identified. In order to better understand the molecular mechanism of Cf-12 gene resistance response, RNA-Seq was used to analyze the transcriptome changes at three different stages of C. fulvum infection (0, 4, and 8 days post infection [dpi]). A total of 9100 differentially expressed genes (DEGs) between 4 and 0 dpi, 8643 DEGs between 8 and 0 dpi and 2547 DEGs between 8 and 4 dpi were identified. In addition, we found that 736 DEGs shared among the above three groups, suggesting the presence of a common core of DEGs in response to C. fulvum infection. These DEGs were significantly enriched in defense-signaling pathways such as the calcium dependent protein kinases pathway and the jasmonic acid signaling pathway. Additionally, we found that many transcription factor genes were among the DEGs, indicating that transcription factors play an important role in C. fulvum defense response. Our study provides new insight on the molecular mechanism of Cf resistance to C. fulvum, especially the unique features of Cf-12 in responding to C. fulvum infection.

Physiological and RNA-seq analyses provide insights into the response mechanism of the Cf-10 -mediated resistance to Cladosporium fulvum infection in tomato

Key messageBased on the physiological and RNA-seq analysis, some progress has been made in elucidating the Cf-10-mediated resistance responses to C. fulvum infection in tomato. GO and KEGG enrichment analysis revealed that the DEGs were significantly associated with defense-signaling pathways like oxidation-reduction processes, oxidoreductase activity and plant hormone signal transduction.AbstractLeaf mold, caused by the fungus Cladosporium fulvum, is one of the most common diseases affecting tomatoes worldwide. Cf series genes including Cf-2, Cf-4, Cf-5, Cf-9 and Cf-10 play very important roles in resisting tomato leaf mold. Understanding the molecular mechanism of Cf gene-mediated resistance is thus the key to facilitating genetic engineering of resistance to C. fulvum infection. Progress has been made in elucidating two Cf genes, Cf -19 and Cf -12, and how they mediate resistance responses to C. fulvum infection in tomato. However, the mechanism of the Cf-10- mediated resistance response is still unclear. In the present study, RNA-seq was used to analyze changes in the transcriptome at different stages of C. fulvum infection. A total of 2,242 differentially expressed genes (DEGs) responsive to C. fulvum between 0 and 16 days post infection (dpi) were identified, including 1,501 upregulated and 741 downregulated genes. The majority of DEGs were associated with defense-signaling pathways including oxidation–reduction processes, oxidoreductase activity and plant hormone signal transduction. Four DEGs associated with plant-pathogen interaction were uniquely activated in Cf-10 tomato and validated by qRT-PCR. In addition, physiological indicators including reactive oxygen species (ROS), superoxide dismutase (SOD), catalase (CAT) and peroxidase (POD) were measured at 0–21 dpi, and hormone expression [Jasmonic acid (JA) and salicylic acid (SA)] was estimated at 0 and 16 dpi to elucidate the mechanism of the Cf-10-mediated resistance response. C. fulvum infection induced the activities of POD, CAT and SOD, and decreased ROS levels. JA was determined to participate in the resistance response to C. fulvum during the initial infection period. The results of this study provide accountable evidence for the physiological and transcriptional regulation of the Cf-10-mediated resistance response to C. fulvum infection, facilitating further understanding of the molecular mechanism of Cf-10-mediated resistance to C. fulvum infection.

Mapping and screening of the tomato Stemphylium lycopersici resistance gene, Sm , based on bulked segregant analysis in combination with genome resequencing

BackgroundTomato gray leaf spot disease caused by Stemphylium lycopersici (S. lycopersici) is considered one of the major diseases of cultivated tomatoes. The only S. lycopersici resistance gene, Sm, was derived from the wild tomato species S. pimpinellifolium. Sm has been identified as an effective source of gray leaf spot resistance in tomatoes and has been mapped to tomato chromosome 11. In this study, the first bulked segregant analysis (BSA) combined with genome resequencing for the mapping and screening of the Sm candidate gene was performed.ResultsBased on the resequencing results, we identified 50,968 Diff-markers, most of which were distributed on chromosome 11. A total of 37 genes were located in the interval of 0.26-Mb. The gene loci of resistant and susceptible lines were sequenced successfully using PCR products. The relative expression levels of candidate genes in resistant and susceptible lines were confirmed via qRT-PCR, Solyc11g011870.1.1 and Solyc11g011880.1.1 were identified through qRT-PCR. A marker, D5, which was cosegregated with the resistant locus, was identified according to the mutation of the Solyc11g011880.1.1 trait in the resistant line.ConclusionsThe Sm gene was mapped to the short arm of chromosome 11. The candidate genes Solyc11g011870.1.1 and Solyc11g011880.1.1 displayed expression patterns related to the resistance response. This study will be valuable for Sm cloning and Sm gene breeding in tomato.