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Transcription profiling of fertilization and early seed development events in a solanaceous species using a 7.7K cDNA microarray from Solanum chacoense ovules

ABSTRACT: To provide a broad analysis of gene expression changes in developing embryos from a solanaceous species, we produced amplicon-derived microarrays with 7741 ESTs isolated from Solanum chacoense ovules bearing embryos from all developmental stages. Our aims were to: 1) identify genes expressed in a tissue-specific and temporal-specific manner; 2) define clusters of genes showing similar patterns of spatial and temporal expression; and 3) identify stage-specific or transition-specific candidate genes for further functional genomic analyses.We analyzed gene expression during S. chacoense embryogenesis in a series of experiments with probes derived from ovules isolated before and after fertilization (from 0 to 22 days after pollination), and from leaves, anthers, and styles. From the 6374 unigenes present in our array, 1024 genes were differentially expressed (≥ ±2 fold change, p value ≤ 0.01) in fertilized ovules and only limited expression overlap was observed between these genes and the genes expressed in the other tissues tested, with more than three quarters of the fertilization-regulated genes specifically or predominantly expressed in ovules. During embryogenesis three major expression profiles corresponding to early, middle and late stages of embryo development were identified. From the early and middle stages, a large number of genes corresponding to cell cycle, DNA processing, signal transduction, and transcriptional regulation were found. Defense and stress response-related genes were found in all stages of embryo development. Protein biosynthesis genes, genes coding for ribosomal proteins and other components of the translation machinery were highly expressed in embryos during the early stage. Genes for protein degradation were overrepresented later in the middle and late stages of embryo development. As expected, storage protein transcripts accumulated predominantly in the late stage of embryo development. Our analysis provides the first study in a solanaceous species of the transcriptional program that takes place during the early phases of plant reproductive development, including all embryogenesis steps during a comprehensive time-course. Our comparative expression profiling strategy between fertilized and unfertilized ovules identified a subset of genes specifically or predominantly expressed in ovules while a closer analysis between each consecutive time points allowed the identification of a subset of stage-specific and transition-specific genes.

ORGANISM(S): Solanum chacoense

PROVIDER: GSE21552 | GEO | 2010/05/13

SECONDARY ACCESSION(S): PRJNA126221

REPOSITORIES: GEO

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Project description:To provide a broad analysis of gene expression changes in developing embryos from a solanaceous species, we produced amplicon-derived microarrays with 7741 ESTs isolated from Solanum chacoense ovules bearing embryos from all developmental stages. Our aims were to: 1) identify genes expressed in a tissue-specific and temporal-specific manner; 2) define clusters of genes showing similar patterns of spatial and temporal expression; and 3) identify stage-specific or transition-specific candidate genes for further functional genomic analyses.We analyzed gene expression during S. chacoense embryogenesis in a series of experiments with probes derived from ovules isolated before and after fertilization (from 0 to 22 days after pollination), and from leaves, anthers, and styles. From the 6374 unigenes present in our array, 1024 genes were differentially expressed (? ±2 fold change, p value ? 0.01) in fertilized ovules and only limited expression overlap was observed between these genes and the genes expressed in the other tissues tested, with more than three quarters of the fertilization-regulated genes specifically or predominantly expressed in ovules. During embryogenesis three major expression profiles corresponding to early, middle and late stages of embryo development were identified. From the early and middle stages, a large number of genes corresponding to cell cycle, DNA processing, signal transduction, and transcriptional regulation were found. Defense and stress response-related genes were found in all stages of embryo development. Protein biosynthesis genes, genes coding for ribosomal proteins and other components of the translation machinery were highly expressed in embryos during the early stage. Genes for protein degradation were overrepresented later in the middle and late stages of embryo development. As expected, storage protein transcripts accumulated predominantly in the late stage of embryo development. Our analysis provides the first study in a solanaceous species of the transcriptional program that takes place during the early phases of plant reproductive development, including all embryogenesis steps during a comprehensive time-course. Our comparative expression profiling strategy between fertilized and unfertilized ovules identified a subset of genes specifically or predominantly expressed in ovules while a closer analysis between each consecutive time points allowed the identification of a subset of stage-specific and transition-specific genes. We produced amplicon-derived microarrays with 7741 ESTs isolated from Solanum chacoense ovules bearing embryos from all developmental stages. To monitor the expression pattern from genes involved in fertilization and embryogenesis processes, flowers were hand-pollinated and ovules were isolated every two days during a 22 days period after pollination. Four independent biological replicates were produced from each time points. In addition, to isolate genes specifically or predominantly expressed in ovules, four biological replicates of leaf, anther, and style tissue mRNA preparations were individually hybridized against unfertilized ovule mRNAs and compared with the data obtained from unfertilized and fertilized ovules at various time points after pollination. To estimate reproducibility and to produce control data for statistical analysis, a large number of unfertilized ovules were isolated and separated between seven independent control groups. RNA from randomly selected pairs of control was hybridized on six microarrays. 6374 unigenes present in our array in duplicate.

Project description:Background. Ovary culture has been a useful way to generate double haploid (DH) plant in cucumber (Cucumis sativus L.). However, the rate of embryo induction and the ability for induced embryo to grow into normal embryo are quite low. Moreover, the s mechanism of cucumber embryogenesis remains ambiguous. In this study, the molecular basis for cucumber embryogenesis was explored to set up basis for a more efficient ovary culture method. Differentially expressed genes during embryogenesis process, including the early stages of embryo formation, embryo maturation and shoot formation, were investigated using transcriptomic sequencing. Methods. Based on the cytological observation of cucumber ovary culture, the ovary culture can be divided into three stages:early embryo development, embryo maturation (from pre-embryos to cotyledon embryos) and the shoot formation stage. six key time points were selected for transcriptome sequencing and analysis. Results. We firstly conducted cytological observations which suggest that cell enlargement is the symbol for gametophytes to switch to sporophyte development pathway during early embryogenesis stage. In this stage, RNA-seq revealed 3468 up-regulated genes, including hormone signal transduction genes, hormone response genes and stress-induced genes. The reported embryogenesis-related genes BBM, HSP90 and AGL were also actively expressed during this stage. The total 480 genes that function in protein complex binding, microtubule binding, tetrapyrrole binding, tubulin binding and other microtubule activities were continuously up-regulated during the embryo maturation stage, indicating that the cytoskeleton structure was continuously being built and maintained by the action of microtubule-binding proteins and enzyme modification during embryo development. In shoot formation stage, 1383 genes were up-regulated, which were mainly enriched in phenylpropanoid biosynthesis, plant hormone signal transduction, phenylalanine metabolism, and starch and sucrose metabolism. The shoot formation stage might be regulated by 6 transcription factors that contained a B3 domain, 9 genes in the AP2/ERF family and 2 genes encoded WUS homologous domain proteins. Conclusions. These findings offer a valuable framework for explaining the transcriptional regulatory mechanism underlying embryogenesis during cucumber ovary culture.

Project description:The significant morphological differences observed during embryo development in angiosperms and gymnosperms are expected to be the result of a differential control of gene expression. We used a loblolly pine (Pinus taeda) cDNA microarray to analyze global transcriptional changes along zygotic embryogenesis in maritime pine (Pinus pinaster). A time-course analysis of the data obtained from the five embryo developmental groups used in this study led to the identification of 4,645 genes whose expression varied along P. pinaster embryogenesis. These transcripts were clustered into six distinct expression profiles. The grouping of these profiles in early, mid-embryogenesis and embryo maturation, according to the developmental period where most of the sequences were up-regulated, evidenced that characteristic transcriptional changes are associated to each developmental period. The application of a cut-off value of 1.95-fold change led to the identification of 1,838 differentially expressed transcripts that were categorized by biological process. Metabolism, interaction with the environment, oxidation-reduction and transport were some of the most represented categories. During early embryogenesis genes putatively involved in phytohormone-mediated signaling were identified, whereas in middle stages the overrepresented genes could be associated with cotyledon formation and induction of somatic embryogenesis. Genes associated with the synthesis of storage products were up-regulated in the latest stages of pine embryo development. It was also during this developmental period that the largest number of sequences putatively encoding transcription factors was identified. Pine immature zygotic embryos were pooled into five different developmental groups according to the collection date (Day0; Day5; Day11; Day15 and Day25). A common reference design was used. Total RNA was extracted from each sample. Three extractions were prepared per sample (biological replicates). A defined ammount from each RNA sample was pooled to use as reference. Messenger RNA was amplified for both test samples and reference. These were hybridized together in two separate experiments (technical replicates). In total, 30 slides were analyzed.

Project description:Long non-coding RNAs (lncRNAs) comprise a diverse class of transcripts that structurally resemble mRNAs but do not encode proteins. Recent genome-wide studies in human and mouse have annotated lncRNAs expressed in cell lines and adult tissues, but a systematic analysis of lncRNAs expressed during vertebrate embryogenesis has been elusive. To identify lncRNAs with potential functions in vertebrate embryogenesis, we performed a time series of RNA-Seq experiments at eight stages during early zebrafish development. We reconstructed 56,535 high-confidence transcripts in 28,912 loci, recovering the vast majority of expressed RefSeq transcripts, while identifying thousands of novel isoforms and expressed loci. We defined a stringent set of 1,133 non-coding multi-exonic transcripts expressed during embryogenesis. These include long intergenic ncRNAs (lincRNAs), intronic overlapping lncRNAs, exonic antisense overlapping lncRNAs, and precursors for small RNAs (sRNAs). Zebrafish lncRNAs share many of the characteristics of their mammalian counterparts: relatively short length, low exon number, low expression, and conservation levels comparable to introns. Subsets of lncRNAs carry chromatin signatures characteristic of genes with developmental functions. The temporal expression profile of lncRNAs revealed two novel properties: lncRNAs are expressed in narrower time windows than protein-coding genes and are specifically enriched in early-stage embryos. In addition, several lncRNAs show tissue-specific expression and distinct subcellular localization patterns. Integrative computational analyses associated individual lncRNAs with specific pathways and functions, ranging from cell cycle regulation to morphogenesis. Our study provides the first comprehensive identification of lncRNAs in a vertebrate embryo and forms the foundation for future genetic, genomic and evolutionary studies. RNA-Seq for 8 zebrafish developmental stages, 2 lanes for each stage (3 for shield).

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Transcription profiling of fertilization and early seed development events in a solanaceous species using a 7.7K cDNA microarray from Solanum chacoense ovules

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