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. 2014 Jan 4:15:9.
doi: 10.1186/1471-2164-15-9.

RNA-Seq derived identification of differential transcription in the chrysanthemum leaf following inoculation with Alternaria tenuissima

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RNA-Seq derived identification of differential transcription in the chrysanthemum leaf following inoculation with Alternaria tenuissima

Huiyun Li et al. BMC Genomics. .

Abstract

Background: A major production constraint on the important ornamental species chrysanthemum is black spot which is caused by the necrotrophic fungus Alternaria tenuissima. The molecular basis of host resistance to A. tenuissima has not been studied as yet in any detail. Here, high throughput sequencing was taken to characterize the transcriptomic response of the chrysanthemum leaf to A. tenuissima inoculation.

Results: The transcriptomic data was acquired using RNA-Seq technology, based on the Illumina HiSeq™ 2000 platform. Four different libraries derived from two sets of leaves harvested from either inoculated or mock-inoculated plants were characterized. Over seven million clean reads were generated from each library, each corresponding to a coverage of >350,000 nt. About 70% of the reads could be mapped to a set of chrysanthemum unigenes. Read frequency was used as a measure of transcript abundance and therefore as an identifier of differential transcription in the four libraries. The differentially transcribed genes identified were involved in photosynthesis, pathogen recognition, reactive oxygen species generation, cell wall modification and phytohormone signalling; in addition, a number of varied transcription factors were identified. A selection of 23 of the genes was transcription-profiled using quantitative RT-PCR to validate the RNA-Seq output.

Conclusions: A substantial body of chrysanthemum transcriptomic sequence was generated, which led to a number of insights into the molecular basis of the host response to A. tenuissima infection. Although most of the differentially transcribed genes were up-regulated by the presence of the pathogen, those involved in photosynthesis were down-regulated.

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Figures

Figure 1
Figure 1
The source of RNA libraries prepared from mock- and pathogen-inoculated chrysanthemum plants. A and B: mock inoculation, C and D: inoculation with an A. tenuissima spore suspension. A, C: The first and fourth true leaves without any treatment. B: the second and third mock-inoculated true leaves. D: the second and third pathogen-inoculated true leaves. Leaves were harvested at 0 h, 6 h, 24 h, 48 h and 72 h after inoculation.
Figure 2
Figure 2
Sequencing saturation analysis of each library. Sequencing saturation in the four libraries of A, B, C and D. The number of different genes detected rose as the read number was increased.
Figure 3
Figure 3
Gene Ontology (GO) classifications of DT genes. DT genes were annotated in three categories: biological process, cellular component and molecular function. Y-axis (right) represents the number of DT genes in each category; Y-axis (left) represents the percentage of a specific category of DT genes within that main category. Panels A, B, C and D (left) represents DT genes in the contrast between library A (leaf 1 and 4 of plants mock-inoculated on leaf 2 and 3) and B (mock-treated leaf 2 and 3) (A-VS-B) (right) , library A and C (leaf 1 and 4 of plants inoculated by the pathogen on leaf 2 and 3) (A-VS-C) (right), library B and D (pathogen-infected leaf 2 and 3) (B-VS-D) (right), library C and D (C-VS-D) (right), respectively.
Figure 4
Figure 4
Distribution of gene coverage analysis of each library. Distribution of gene coverage in libraries A, B, C and D.
Figure 5
Figure 5
The number of DT unigenes identified in each library contrast.

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