Open Access Research article

Deep mRNA sequencing reveals stage-specific transcriptome alterations during microsclerotia development in the smoke tree vascular wilt pathogen, Verticillium dahliae

Dianguang Xiong1, Yonglin Wang1, Jie Ma2, Steven J Klosterman3, Shuxiao Xiao1 and Chengming Tian1*

Author Affiliations

1 The Key Laboratory for Silviculture and Conservation of Ministry of Education, College of Forestry, Beijing Forestry University, Beijing, China

2 School of Information Science and Technology, Beijing Forestry University, Beijing, China

3 United States Department of Agriculture-Agricultural Research Service, Salinas, CA, USA

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BMC Genomics 2014, 15:324  doi:10.1186/1471-2164-15-324

Published: 1 May 2014

Additional files

Additional file 1: Figure S1:

Overview of the RNA-Seq data. All of the developmental stages share similar sets of mapped reads. The number of reads produced from the Illumina platform (blue bar); the number of reads mapped to the reference genome within 2 bp mismatch (red bar); the number of reads perfectly mapped to the reference genome (green bar); the number of reads mapped to the annotated genes (purple bar).

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Additional file 2: Figure S2:

Validation of RNA-Seq expression patterns. The RT-qPCR results of the selected genes show similar expression patterns to those detected by RNA-Seq.

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Additional file 3: Table S1:

GO enrichment analysis of genes in 18 clusters (p_value < 0.01).

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Additional file 4: Figure S3:

Expression profiles of MAPK cascades. Heatmap showed that four types of MAPK cascades during MS formation. Expression pattern of genes encoding high osmolarity (HOG1) pathway were clustered together. The non-LS HOG1-MAPK maintained the highest expression value compared to other MAPKs.

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Additional file 5: Figure S4:

Expression profiles of small secreted proteins (length ≤300aa). Expression profiles of those genes encoding small secreted proteins with at least 50-fold up-regulated during MS formation compared with the CO stage. The levels of expression represent log2 FPKM Value +1. * represents genes with cysteine residues <4.

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Additional file 6: Table S2:

List of genes involved in metabolism, signal pathway and identified transcription factor.

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Additional file 7: Figure S5:

Functional categorization of genes differentially expressed (up- or down-regulated) during microsclerotia development. (A, C, E) Intersection of MS1-4 stages revealed 600 significantly up-regulated genes vs CO stage, and functional categorization of these genes. (B, D, F) Intersection of MS1-4 stages revealed 124 significantly down-regulated genes vs CO stage, and functional categorization of these genes.

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Additional file 8: Table S3:

Significantly regulated genes during microsclerotia formation vs CO stage (p_value < 0.05).

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Additional file 9: Figure S6:

Genes involved in protein metabolic processes and autophagy. A. Genes involved in protein metabolic processes by functional categorization; genes labeled with asterisks are the subunits associated with proteasome formation. B. Heatmap representation of genes involved in autophagy processes. Heatmap shows levels of transcripts abundance; relative levels of expression are presented by moderated log2 ratio of transcript abundance in MS developmental stages relative to the CO stage.

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Additional file 10: Figure S7:

Phylogenetic analysis and expression profile of bZIP transcription factors of V. dahliae. The full-length amino acid sequences of bZIP transcription factors of V. dahliae strain VdLs.17 were aligned using Clustal X and the phylogenetic tree was constructed using Mega 5.0 using the maximum-likelihood method with 1000 replicates. Domain structures are drawn to represent their relative positions. The black solid line represents the corresponding protein and its length. The different-colored boxes represent different domains and their positions in each protein predicted by Sanger Pfam program (http://pfam.xfam.org/ webcite) and the yellow ovals represent bZIP domains. The red box indicates an independent cluster of bZIP transcription factors genes, which contain two specific motifs represented at the bottom. The right panel represents heat maps showing the expression pattern of bZIP transcription factors of V. dahliae.

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Additional file 11: Figure S8:

Phylogenetic analysis of bZIP transcription factors of V. dahliae and other fungi. The amino acid sequences of bZIP transcriptional factors of V. dahliae strain VdLs.17, and other fungi were aligned using Clustal X and the phylogenetic tree was constructed using Mega 5.0 using the maximum-likelihood method with 1000 replicates. Fungal species are Sc, Saccharomyces cerevisiae; Vd, Verticillium dahliae; Va, Verticillium alfalfae (formerly V. albo-atrum); Fv, Fusarium verticillioides; Fo, Fusarium oxysporum; Fg, Fusarium graminearum; Mg, Magnaporthe oryzae; Nc, Neurospora crassa. Pa, Podospora anserine; Hj, Hypocrea jecorina.

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Additional file 12: Figure S9:

Expression patterns of genes involved in glycometabolism. Genes involved in carbohydrate metabolism including Glycolysis/Gluconeogensis, Glyoxylate cycle, TCA cycle, Pentose phosphate pathway. Heatmap shows levels of transcripts abundance; Level of expression are presented by moderated log2 ratio of transcript abundance vs CO stage.

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Additional file 13: Table S4:

List of genes undergoing AS events.

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Additional file 14: Table S5:

Statistics of the AS events genes.

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Additional file 15: Figure S10:

RT-PCR validation of three genes undergo RI events. Primers (black arrows) were designed spanning intron region. gDNA represents genomic DNA. cDNA represents complementary DNA.

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Additional file 16: Table S6:

Gene expression value of all annotated genes.

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Additional file 17: Table S7:

Quality of Total RNA by Agilent 2000.

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Additional file 18: Table S8:

RT-PCR primers used in this study.

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