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Single cell RNA sequencing of peanut-responsive T cells

ABSTRACT: Peanut-responsive T cells from peanut allergic subjects were identified and selected based on CD154 expression after stimulation of peripheral blood mononuclear cells with crude peanut extract for 18h. As controls, polyclonally activated CD4+ T cells from peanut allergic subjects were selected. Additional controls included CD4+CD25+CD127- Tregs from peanut allergic or healthy controls. Single cells were obtained using the C1 system from Fluidigm, and a barcoded library constructed. Sequencing (Illumina) was performed using 100 nt paired end reads. Data on a total of 431 cells was available. The goal of the study was to understand the heterogeneity of the peanut-specific T cell response.

ORGANISM(S): Homo sapiens

PROVIDER: GSE98852 | GEO | 2017/12/01

SECONDARY ACCESSION(S): PRJNA386454

REPOSITORIES: GEO

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Project description:To gain insights into molecular mechanisms of tolerance to heat stress, we conducted a transcript profiling experiment to identify heat-responsive genes in contrasting peanut mini core accessions, either un-acclimated or acclimated to heat stress. Plants at reproductive stage were exposed to 28 °C (control), 45 °C for 15 d (un-acclimated) or 45 °C for 1 d followed by 7 d recovery and 15 d stress (acclimated). Two contrasting genotypes showing diverse response to stress were selected based on a bioassay involving chlorophyll fluorescence yield under elevated respiratory demand and membrane thermostability. Transcript profiling was performed using 8 x 15k custom oligo microarrays containing 15k peanut EST sequences. Gene enrichment analysis was performed using Blast2GO program and genes with homology to known proteins were categorized into detailed molecular functional groups. Majority of stress-responsive genes assigned to KEGG pathways belonged to starch, sucrose and galactose metabolism followed by aminoacid metabolism, and secondary metabolite biosynthesis. Differentially expressed transcripts from samples were validated in the samples from second year by quantitative real-time PCR. Transcripts of eight genes involved in terpenoid and flavanoid biosynthesis were induced after second and seventh day, respectively in leaves under heat stress. Metabolite analysis confirmed increases in metabolites of selected pathways under heat stress. The heat up-regulated genes in tolerant COC041 mini-core accession are potential candidate genes for engineering stress-tolerant peanuts and unraveling molecular mechanisms of peanut adaptation to heat stress. We used Agilent peanut microarrays to identify putative heat stress-responsive genes. Acclimated leaf tissues of the peanut genotypes COC041 (tolerant) and COC166 (susceptible) were used in the study. Three replications of microarray experiments were carried out by hybridizing the cRNA from different time points and stress conditions in a loop design on 8 x 15k microarray.

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Single cell RNA sequencing of peanut-responsive T cells

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