Dataset Information

Agrobacterium tumefaciens-mediated transformation and expression of GFP in Ascochyta lentis to characterize ascochyta blight disease progression in lentil.

ABSTRACT: The plant immune system is made up of a complex response network that involves several lines of defense to fight invading pathogens. Fungal plant pathogens on the other hand, have evolved a range of ways to infect their host. The interaction between Ascochyta lentis and two lentil genotypes was explored to investigate the progression of ascochyta blight (AB) in lentils. In this study, we developed an Agrobacterium tumefaciens-mediated transformation system for A. lentis by constructing a new binary vector, pATMT-GpdGFP, for the constitutive expression of green fluorescent protein (EGFP). Green fluorescence was used as a highly efficient vital marker to study the developmental changes in A. lentis during AB disease progression on the susceptible and resistant lentil accessions, ILL6002 and ILL7537, respectively. The initial infection stages were similar in both the resistant and susceptible accessions where A. lentis uses infection structures such as germ tubes and appressoria to gain entry into the host while the host uses defense mechanisms to prevent pathogen entry. Penetration was observed at the junctions between neighbouring epidermal cells and occasionally, through the stomata. The pathogen attempted to penetrate and colonize ILL7537, but further fungal advancement appeared to be halted, and A. lentis did not enter the mesophyll. Successful entry and colonization of ILL6002 coincided with structural changes in A. lentis and the onset of necrotic lesions 5-7 days post inoculation. Once inside the leaf, A. lentis continued to grow, colonizing all parts of the leaf followed by plant cell collapse. Pycnidia-bearing spores appeared 14 days post inoculation, which marks the completion of the infection cycle. The use of fluorescent proteins in plant pathogenic fungi together with confocal laser scanning microscopy, provide a valuable tool to study the intracellular dynamics, colonization strategy and infection mechanisms during plant-pathogen interaction.

SUBMITTER: Henares BM

PROVIDER: S-EPMC6812748 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Agrobacterium tumefaciens-mediated transformation and expression of GFP in Ascochyta lentis to characterize ascochyta blight disease progression in lentil.

Henares Bernadette M BM Debler Johannes W JW Farfan-Caceres Lina M LM Grime Christina R CR Lee Robert C RC

PloS one 20191024 10

The plant immune system is made up of a complex response network that involves several lines of defense to fight invading pathogens. Fungal plant pathogens on the other hand, have evolved a range of ways to infect their host. The interaction between Ascochyta lentis and two lentil genotypes was explored to investigate the progression of ascochyta blight (AB) in lentils. In this study, we developed an Agrobacterium tumefaciens-mediated transformation system for A. lentis by constructing a new bin ...[more]

PMID: 31647840

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Project description:Ascochyta blight of lentil is an important fungal disease in many lentil-producing regions of the world causing major yield and grain quality losses. Quick shifts in aggressiveness of the population of the causal agent Ascochyta lentis mandates developing germplasm with novel and durable resistance. In the absence of complete resistance, lentil genotypes CDC Robin and 964a-46 have frequently been used as sources of partial resistance to ascochyta blight and carry non-allelic ascochyta blight resistance genes. RNA-seq analysis was conducted to identify differences in the transcriptome of CDC Robin, 964a-46 and the susceptible check Eston after inoculation with A. lentis. Candidate defense genes differentially expressed among the genotypes had hypothetical functions in various layers of plant defense, including pathogen recognition, phytohormone signaling pathways and downstream defense responses. CDC Robin and 964a-46 activated cell surface receptors (e.g. receptor like kinases) tentatively associated with pathogen-associated molecular patterns (PAMP) recognition and nucleotide-binding site leucine-rich repeat (NBS-LRR) receptors associated with intracellular effector recognition upon A. lentis infection, and differed in their activation of salicylic acid, abscisic acid and jasmonic acid / ethylene signal transduction pathways. These differences were reflected in the differential expression of downstream defense responses such as pathogenesis-related proteins, and genes associated with the induction of cell death and cell-wall reinforcement. A significant correlation between expression levels of a selection of genes based on quantitative real-time PCR and their expression levels estimated through RNA-seq demonstrated the technical and analytical accuracy of RNA-seq for identification of genes differentially expressed among genotypes. The presence of different resistance mechanisms in 964a-46 and CDC Robin indicates their value for pyramiding gene leading to more durable resistance to ascochyta blight.

Project description:Lentil (Lens culinaris Medik.) is a self-pollinating, diploid, annual, cool-season, food legume crop that is cultivated throughout the world. Ascochyta blight (AB), caused by Ascochyta lentis Vassilievsky, is an economically important and widespread disease of lentil. Development of cultivars with high levels of durable resistance provides an environmentally acceptable and economically feasible method for AB control. A detailed understanding of the genetic basis of AB resistance is hence highly desirable, in order to obtain insight into the number and influence of resistance genes. Genetic linkage maps based on single nucleotide polymorphisms (SNP) and simple sequence repeat (SSR) markers have been developed from three recombinant inbred line (RIL) populations. The IH × NF map contained 460 loci across 1461.6 cM, while the IH × DIG map contained 329 loci across 1302.5 cM and the third map, NF × DIG contained 330 loci across 1914.1 cM. Data from these maps were combined with a map from a previously published study through use of bridging markers to generate a consensus linkage map containing 689 loci distributed across seven linkage groups (LGs), with a cumulative length of 2429.61 cM at an average density of one marker per 3.5 cM. Trait dissection of AB resistance was performed for the RIL populations, identifying totals of two and three quantitative trait loci (QTLs) explaining 52 and 69% of phenotypic variation for resistance to infection in the IH × DIG and IH × NF populations, respectively. Presence of common markers in the vicinity of the AB_IH1- and AB_IH2.1/AB_IH2.2-containing regions on both maps supports the inference that a common genomic region is responsible for conferring resistance and is associated with the resistant parent, Indianhead. The third QTL was derived from Northfield. Evaluation of markers associated with AB resistance across a diverse lentil germplasm panel revealed that the identity of alleles associated with AB_IH1 predicted the phenotypic responses with high levels of accuracy (~86%), and therefore have the potential to be widely adopted in lentil breeding programs. The availability of RIL-based maps, a consensus map, and validated markers linked to AB resistance provide important resources for lentil improvement.

Dataset Information

Agrobacterium tumefaciens-mediated transformation and expression of GFP in Ascochyta lentis to characterize ascochyta blight disease progression in lentil.

Publications

Agrobacterium tumefaciens-mediated transformation and expression of GFP in Ascochyta lentis to characterize ascochyta blight disease progression in lentil.

Similar Datasets

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