Background Rice is an important food source for humans worldwide. Because of its nutritional and agricultural significance, a number of studies addressed various aspects of rice grain development and grain filling. Nevertheless, the molecular processes underlying grain filling and development, and in particular the contributions of different grain tissues to these processes, are not understood. Main Text Using RNA-sequencing, we profiled gene expression activity in grain tissues comprised of cross cells (CC), the nucellar epidermis (NE), ovular vascular trace (OVT), endosperm (EN) and the aleurone layer (AL). These tissues were dissected using laser capture microdissection (LCM) at three distinct grain development stages. The mRNA expression datasets offer comprehensive and new insights into the gene expression patterns in different rice grain tissues and their contributions to grain development. Comparative analysis of the different tissues revealed their similar and/or unique functions, as well as the spatio-temporal regulation of common and tissue-specific genes. The expression patterns of genes encoding hormones and transporters indicate an important role of the OVT tissue in metabolite transport during grain development. Gene co-expression network prediction on OVT-specific genes identified several distinct and common development-specific transcription factors. Further analysis of enriched DNA sequence motifs proximal to OVT-specific genes revealed known and novel DNA sequence motifs relevant to rice grain development. Conclusion Together, the dataset of gene expression in rice grain tissues is a novel and useful resource for further work to dissect the molecular and metabolic processes during rice grain development.

Microarrays are routine tools for transcript profiling, and genomic tiling arrays such as the Arabidopsis AGRONOMICS1 arrays have been found to be highly suitable for such experiments because changes in genome annotation can be easily integrated at the data analysis level. In a transcript profiling experiment, RNA labeling is a critical step, most often initiated by oligo-dT-primed reverse transcription. Although this has been found to be a robust and reliable method, very long transcripts or non-polyadenylated transcripts might be labeled inefficiently. In this study, we first provide data handling methods to analyze AGRONOMICS1 tiling microarrays based on the TAIR10 genome annotation. Second, we describe methods to easily quantify antisense transcripts on such tiling arrays. Third, we test a random-primed RNA labeling method, and find that on AGRONOMICS1 arrays this method has similar general performance as the conventional oligo-dT-primed method. In contrast to the latter, however, the former works considerably better for long transcripts and for non-polyadenylated transcripts such as found in mitochondria and plastids. We propose that researchers interested in organelle function use the random-primed method to unleash the full potential of genomic tiling arrays.

Background Rice is an important food source for humans worldwide. Because of its nutritional and agricultural significance, a number of studies addressed various aspects of rice grain development and grain filling. Nevertheless, the molecular processes underlying grain filling and development, and in particular in different contributions of grain tissues to these process, are not understood.Main text Using RNA-sequencing, we profiled gene expression activity in grain tissues comprised of cross cells (CC), the nucellar epidermis (NE), ovular vascular trace (OVT), endosperm (EN) and the aleurone layer (AL). These tissues were dissected using laser capture microdissection (LCM) at three distinct grain development stages. The mRNA expression datasets offer comprehensive and new insights into the gene expression patterns in different rice grain tissues and their contributions to grain development. Comparative analysis of the different tissues revealed their similar and/or unique functions, as well as the spatio-temporal regulation of common and tissue-specific genes. The expression patterns of genes encoding hormones and transporters indicate an important role of the OVT tissue in metabolite transport during grain development. Gene co-expression network prediction on OVT-specific genes identified several distinct and common development-specific transcription factors. Further analysis of enriched DNA sequence motifs proximal to OVT-specific genes revealed known and novel DNA sequence motifs relevant to rice grain development.Conclusion Together, the dataset of gene expression in rice grain tissues is a novel and useful resource for further work to dissect the molecular and metabolic processes during rice grain development.Gene expression was analyzed in five sub-regions of the developing rice grain comprising the CC, NE, OVT, EN and AL tissues that were collected by laser-capture micro dissection at 4, 8 and 16 DAF. RNA sequence reads were quality checked and over 20,000 genes were identified with statistically significant expression (reads higher than 10, p < 0.001). Genes with expression specific to each subregion and grain development stages were compared using hierarchical clustering and principal component analysis (PCA). The biological replicates from individual sub-regions clustered together and PCA collectively identified 79% variance in our dataset (Fig. 1, Fig. S1). The dataset was further analyzed to separate genes for their tissue-and stage-specific expression. The genes with a preferentially higher expression level (with the highest number of reads, p < 0.001) in a particular sub-region or at a particular developmental time point were assigned to be tissue-specific and stagespecific differentially expressed genes (DEGs), respectively. In total, 10,037 DEGs were identified. In order to further characterize the sub-region and stage-specific genes, gene ontology (GO) analysis 5 was conducted on DEGs using PlantGSEA (Yi et al., 2013). Significantly overrepresented GO terms and metabolic pathways (p < 0.01) wer...

Background Rice is an important food source for humans worldwide. Because of its nutritional and agricultural significance, a number of studies addressed various aspects of rice grain development and grain filling. Nevertheless, the molecular processes underlying grain filling and development, and in particular in different contributions of grain tissues to these process, are not understood. Main text Using RNA-sequencing, we profiled gene expression activity in grain tissues comprised of cross cells (CC), the nucellar epidermis (NE), ovular vascular trace (OVT), endosperm (EN) and the aleurone layer (AL). These tissues were dissected using laser capture microdissection (LCM) at three distinct grain development stages. The mRNA expression datasets offer comprehensive and new insights into the gene expression patterns in different rice grain tissues and their contributions to grain development. Comparative analysis of the different tissues revealed their similar and/or unique functions, as well as the spatio-temporal regulation of common and tissue-specific genes. The expression patterns of genes encoding hormones and transporters indicate an important role of the OVT tissue in metabolite transport during grain development. Gene co-expression network prediction on OVT-specific genes identified several distinct and common development-specific transcription factors. Further analysis of enriched DNA sequence motifs proximal to OVT-specific genes revealed known and novel DNA sequence motifs relevant to rice grain development. Conclusion Together, the dataset of gene expression in rice grain tissues is a novel and useful resource for further work to dissect the molecular and metabolic processes during rice grain development.