Project description:The sea-ice dwelling diatom Fragilariopsis Cylindrus was cultured for 4 months under dark or light exposed conditions to mimic the effects of Antarctic winter growth conditions. Cells were harvested periodically and LFQ proteomics used to investigate the molecular mechanisms of dark survival.
Project description:We performed RNA-sequencing experiments to examine the differential regulation of genes in the genome of the Southern Ocean diatom Fragilariopsis cylindrus including diverged alleles. RNA-seq was performed on three replicate samples for each experimental condition. Phytoplankton cells were grown under six different experimental conditions including (1) optimal growth, (2) freezing temperatures, (3) elevated temperature, (4) elevated carbon dioxide concentrations, (5) low iron concentrations and (6) prolonged darkness. Total RNA was extracted using a guanidinium thiocyanate-phenol-chloroform extraction protocol, followed by DNase I treatment and RNA purification (Quiagen). First strand cDNA synthesis was performed using random hexamers. Library preparation was performed using the RNA-seq Sample Prep Kit (Illumina) and sequencing was conducted according to the TruSeq RNA sequencing protocol (Illumina) All samples were sequenced together in one flowcell on one lane.
Project description:The Southern Ocean diatom Fragilariopsis cylindrus was grown under half-saturating concentrations of silicate (0.3 uM) and in the blue (480 - 540 nm) and red (550 - 700 nm) light spectrum to investigate gene expression responses using RNA-seq relative to optimal growth conditions (deposited under accession E-MTAB-5024). RNA-seq was performed on three replicate samples for each experimental condition. Total RNA was extracted using a guanidinium thiocyanate-phenol-chloroform extraction protocol, followed by DNase I treatment and RNA purification (Quiagen). First strand cDNA synthesis was performed using random hexamers. Library preparation was performed using the RNA-seq Sample Prep Kit (Illumina) and sequencing was conducted according to the TruSeq RNA sequencing protocol (Illumina) All samples were sequenced together in one flowcell on one lane.
Project description:RNA-seq transcriptome profiling of the psychrophilic phytoplankton Fragilariopsis cylindrus under environmentally relevant growth limitations
Project description:Background: Marine phytoplankton are responsible for 50% of the CO2 that is fixed annually worldwide and contribute massively to other biogeochemical cycles in the oceans. Diatoms and coccolithophores play a significant role as the base of the marine food web and they sequester carbon due to their ability to form blooms and to biomineralise. To discover the presence and regulation of short non-coding RNAs (sRNAs) in these two important phytoplankton groups, we sequenced short RNA transcriptomes of two diatom species (Thalassiosira pseudonana, Fragilariopsis cylindrus) and validated them by Northern blots along with the coccolithophore Emiliania huxleyi. Results: Despite an exhaustive search, we did not find canonical miRNAs in diatoms. The most prominent classes of sRNAs in diatoms were repeat-associated sRNAs and tRNA-derived sRNAs. The latter were also present in E. huxleyi. tRNA-derived sRNAs in diatoms were induced under important environmental stress conditions (iron and silicate limitation, oxidative stress, alkaline pH), and they were very abundant especially in the polar diatom F. cylindrus (20.7% of all sRNAs) even under optimal growth conditions. Conclusions: This study provides first experimental evidence for the existence of short non-coding RNAs in marine microalgae. Our data suggest that canonical miRNAs are absent from diatoms. However, the group of tRNA-derived sRNAs seems to be very prominent in diatoms and coccolithophores and may be used for acclimation to environmental conditions.
Project description:The genome of the cold-adapted diatom Fragilariopsis cylindrus is characterized by highly diverged haplotypes that intersperse its homozygous genome. Here, we describe how a combination of PacBio DNA and Illumina RNA sequencing can be used to resolve this complex genomic landscape locally into the highly diverged haplotypes, and how to map various environmentally controlled transcripts onto individual haplotypes. We assembled PacBio sequence data with the FALCON assembler and created a haplotype resolved annotation of the assembly using annotations of a Sanger sequenced F. cylindrus genome. RNA-seq datasets from six different growth conditions were used to resolve allele-specifc gene expression in F. cylindrus. This approach enables to study differential expression of alleles in a complex genomic landscape and provides a useful tool to study how diverged haplotypes in diploid organisms are used for adaptation and evolution to highly variable environments.
Project description:RNA-seq transcriptome profiling of the psychrophilic diatom Fragilariopsis cylindrus under six treatments simulating conditions within sea ice
