Life on a sand grain
Ontology highlight
ABSTRACT:
PROVIDER: PRJEB20733 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| ERR2113130.fastq.gz | Fastqsanger.gz | |||
| ERR2113140.fastq.gz | Fastqsanger.gz | |||
| ERR2113131.fastq.gz | Fastqsanger.gz | |||
| ERR2113141.fastq.gz | Fastqsanger.gz | |||
| ERR2113122.fastq.gz | Fastqsanger.gz |
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Project description:Transcriptomic analyses are particularly powerful in research when they are based on detailed underlying knowledge of developmental processes, morphological/anatomical features and biochemical/metabolic processes. Our work is based on our detailed knowledge of grain development and provides an invaluable resource for both Brachypodium grain development specifically and for comparative analyses to other species, including cereal crops. Grain development includes several transition points in addition to being preceded and succeeded by the fundamental developmental switches of fertilization and germination. Such complex process requires a larger sets of genes expressed in a highly-controlled manner. We have selected eight stages of developmental in Brachypodium distachyon encompassing these transitions and conducted comprehensive transcriptomic analyses to generate a valuable resource of the developmental transitions and the distinctive biological processes that are activated and/or repressed during grain development and germination. Comparison of the data generated in this project to other grasses, including the important crop species, and beyond will also shed light on the conservation and/or diversification in gene expression and function in an evolutionary context. Brachypodium distachyon grains (Bd-21 accession) were germinated on moist filter paper after stratification at 4°C for 48h. Five-day old seedlings were transferred to 9cm square pots with a 2:2:1 multipurpose compost: vermiculite: sand mix and grown under controlled environment conditions with a 18h photoperiod at 20°-22°C and light intensity of 180-200 umol/m/s. Tissue samples were collected from eight distinct developmental stages; pre-anthesis ovaries, young grains (1-3 DAA), middle length grains (3-8 DAA), full length grains (8-15 DAA), mature grains (15-20 DAA), mature grains without embryo, germinating grains and seedlings at 3-4 days after germination (Fig.1). Tissues were frozen in liquid nitrogen immediately after collection and were kept at -80°C until further analyses. Total RNA was isolated from frozen tissue samples using the Spectrum Plant Total RNA Kit (Sigma-Aldrich) following the manufacturer’s instructions. Three biological replicates were used for each stage of development. The RNA samples were treated with DNase I (New England Biolabs) in order to eliminate DNA contamination and their concentration was determined using a NanoDrop spectrophotometer (Thermo Fisher Scientific).
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Project description:A laboratory colony of Phlebotomus perniciosus sand flies was maintained. Sand flies were infected with cultured Leishmania infantum promastigotes in stationary phase. Ten infected sand flies were dissected after 5 days and promastigotes within the gut pooled. The cells were immediately washed in PBS once and lysed in TRIzol reagent (Life Technologies). RNA isolation was completed according to the manufacturer's instructions, obtaining 63ng. RNA-seq libraries were generated using the spliced leader sequence for second strand synthesis (Cuypers et al., 2017; Haydock et al., 2015), thus allowing for specific amplification of sequences from L. infantum promastigotes, thus avoiding contamination with material from the sand fly gut. Single-end sequencing was performed in an Illumina HiSeq2500 instrument and data analysis was conducted using bowtie2, samtools, featureCounts and Geneious. The main findings are: i) substantial differences in differential gene expression between sand fly-derived (sfPro) and cultured (acPro) promastigotes; and ii) over-expression of genes involved in metacyclogenesis in sfPro vs. acPro, including gp63 genes, autophagy genes, etc.
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Project description:Crop reproduction is highly sensitive to water-deficit and heat stress. The molecular networks of stress adaptation and grain development in tetraploid wheat (T. turgidum durum) are not well understood. Small RNAs (sRNAs) are important epigenetic regulators connecting the transcriptional and post-transcriptional regulatory networks. This study presents the first multi-omics analysis of the sRNAome, transcriptome and degradome in T. turgidum developing grains, under single and combined water-deficit and heat stress. We identified 690 microRNAs (miRNAs), with 84 being novel, from 118 sRNA libraries. Complete profiles of differentially expressed miRNA (DEMs) specific to genotypes, stress types and different reproductive time-points are provided. The first degradome-seq report for developing durum grains discovered a significant number of new target genes regulated by miRNAs post-transcriptionally. Transcriptome-seq profiled 53,146 T. turgidum genes, with differentially expressed genes (DEGs) enriched in functional categories such as nutrient metabolism, cellular differentiation, transport, reproductive development and hormone transduction pathways. miRNA-mRNA networks that affect grain characteristics such as starch synthesis and protein metabolism were constructed, based on integrated analysis of the three omics. This study provides a substantial amount of novel information on the post-transcriptional networks in T. turgidum grains, which will facilitate innovations for breeding programs aiming to improve crop resilience and grain quality.
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