We describe a method for isolating RNA suitable for high-throughput RNA sequencing (RNA-seq) from small numbers of fluorescently labeled cells isolated from live zebrafish (Danio rerio) embryos without using costly, commercially available columns. This method ensures high cell viability after dissociation and suspension of cells and gives a very high yield of intact RNA. We demonstrate the utility of our new protocol by isolating RNA from fluorescence activated cell sorted (FAC sorted) pineal complex neurons in wild-type and tbx2b knockdown embryos at 24 hours post fertilization. Tbx2b is a transcription factor required for pineal complex formation. We describe a bioinformatics pipeline used to analyze differential expression following high-throughput sequencing and demonstrate the validity of our results using in situ hybridization of differentially expressed transcripts. This protocol brings modern transcriptome analysis to the study of small cell populations in zebrafish. Overall design: Differential expression analysis of mRNA levels in a single time-point (24 hpf) between wild-type and Tbx2b deficient FAC sorted pineal complex cells
Identification of differentially expressed genes during development of the zebrafish pineal complex using RNA sequencing.
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View SamplesAbiotic stresses like drought, salinity, high and low temperature, and submergence are major factors that limit the crop productivity. Hence, identification of genes associated with stress response in crops is a prerequisite for improving their tolerance to adverse environmental conditions. In this study, we have analyzed the expression profiles of three genotypes WT, TaVAP mutant and TaVAPOE plants in Arabidopsis thaliana in col-0 background using microarray technology to identify the genes differentially expressed under control conditions.
Gene encoding vesicle-associated membrane protein-associated protein from Triticum aestivum (TaVAP) confers tolerance to drought stress.
Specimen part
View SamplesNatural killer (NK) cells can be divided into phenotypic subsets based on the expression of receptors that bind self-MHC-I molecules with differing affinities; a concept termed licensing or education. Here we show that NK cell subsets exhibit markedly different migratory, effector, and immunoregulatory functions on dendritic cells and antigen-specific CD8+ T cell responses during influenza and murine cytomegalovirus infections. Shortly after infection, unlicensed NK cells preferentially trafficked to draining lymph nodes and produced GM-CSF, which promoted the expansion and activation of dendritic cells, and ultimately resulted in sustained antigen-specific CD8+ T cell responses. In contrast, licensed NK cells preferentially migrated to infected parenchymal tissues and produced greater levels of interferon- (IFN-). Importantly, human NK cell subsets exhibited similar phenotypic characteristics and patterns of cytokine production. Collectively, our studies demonstrate a critical demarcation between the functions of licensed and unlicensed NK cell subsets, with the former functioning as the classical effector subset in inflamed tissues and the latter as modulators of adaptive immunity helping to prime immune responses in draining lymph nodes.
Licensing delineates helper and effector NK cell subsets during viral infection.
Specimen part
View SamplesRice transgenic plants of the F-box encoding gene, OsFBK1, in the Pusa Basmati 1 (PB1) variety have been found to display differences in the anther and root phenotypes. In order to elucidate changes at the transcriptome level, microarray of the roots of 14-day-old seedlings of over-expresseion (OE) and knock-down (KD) lines along with vector control (VC) and wild type (WT) were carried out.
The OsFBK1 E3 Ligase Subunit Affects Anther and Root Secondary Cell Wall Thickenings by Mediating Turnover of a Cinnamoyl-CoA Reductase.
Specimen part
View SamplesIndica rice seedlings of IR64 variety were grown hydroponically for 7-days in a culture room with a daily photoperiodic cycle of 14h light and 10h dark. Seedlings were incubated in 0.1% dimethyl sulfoxide (control) or 50 micromolar solutions of indole-3-acetic acid (IAA treatment) and benzyl aminopurine (BAP treatment) for 1h and 3h. Equal amounts of 1h and 3h samoles were pooled for each treatment before RNA isolation. The 5 micrograms of each total RNA sample was processed for microarray analysis according to Affymetrix protocol.
Transcript profiling reveals diverse roles of auxin-responsive genes during reproductive development and abiotic stress in rice.
Specimen part
View SamplesRice transgenic plants of the bZIP encoding gene, OsbZIP48, in the Pusa Basmati 1 (PB1) variety have been found to display differences in the total height. In order to elucidate changes at the transcriptome level, microarray of the 10-day-old seedlings of over-expresseion (OE) and knock-down (KD) lines along with vector control (VC) were carried out.
OsbZIP48, a HY5 Transcription Factor Ortholog, Exerts Pleiotropic Effects in Light-Regulated Development.
Age, Specimen part
View Samples10-day-old wild-type and homozygous transgenic Arabidopsis seedlings (overexpressing OsTOP6A3 and OsTOP6B) grown under normal growth conditions were used for total RNA isolation. The 5 micrograms of each total RNA sample was processed for microarray analysis according to Affymetrix protocol.
Overexpression of putative topoisomerase 6 genes from rice confers stress tolerance in transgenic Arabidopsis plants.
Specimen part
View Samples10-day-old wild-type and homozygous transgenic Arabidopsis seedlings (overexpressing rice topoisomerase 6 subunit A1; OsTOP6A1) grown under normal growth conditions were used for total RNA isolation. The 5 micrograms of each total RNA sample was processed for microarray analysis according to Affymetrix protocol.
Constitutive expression of a meiotic recombination protein gene homolog, OsTOP6A1, from rice confers abiotic stress tolerance in transgenic Arabidopsis plants.
Specimen part
View SamplesHomeobox transcription factors are known to regulate plant growth and development. Recently, they have also been implicated in abiotic stress responses. To analyze the role of HD-ZIP I subfamily member, OsHOX24, we constitutively overexpressed it in Arabidopsis. The physiological analyses revealed that overexpression of OsHOX24 gene severely reduced abiotic stress tolerance in transgenic plants as compared to wild-type.
Characterization of Rice Homeobox Genes, OsHOX22 and OsHOX24, and Over-expression of OsHOX24 in Transgenic Arabidopsis Suggest Their Role in Abiotic Stress Response.
Age, Specimen part
View SamplesThis SuperSeries is composed of the SubSeries listed below.
Influence of hyperthyroid conditions on gene expression in extraocular muscles of rats.
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