Genomics,Multiomics

Dataset Information

25

Single-cell RNA sequencing in Trypanosoma brucei


ABSTRACT: Trypanosomes were sorted (0 cells, 1 cell, 50 cells) using a FACSaria III (BD Biosciences; precision: single-cell; nozzle: 100 µm). Forward-scatter area (FCS-A) versus side-scatter area (SSC-A) was used to gate the cells. Trypanosomes were sorted in 48-wells plate (Brand) filled with 2.6 µL of lysis buffer (0.01 µL of RNAse inhibitor (Takara) and 1x Lysis buffer (Takara) in RNAse-free water). Immediately after sorting cells were placed on ice for 5 minutes and stored at -80 °C. 50 and single trypanosomes were prepared using SMART-Seq v4 Ultra Low Input RNA Kit (Takara) using one fourth of reagents volumes compared to the supplier instructions. PCR amplification was performed using 26 cycles using supplier recommendations. cDNA was purified using XP beads (Beckman Coulter) and recovered in 15 µL of elution buffer (Takara). Libraries were quantified using the Qubit Hs Assay (Life Technologies) and the qualities of the libraries were further monitored using a Bioanalyzer (Agilent). Similar to what has been published previously 19, 1 ng of cDNA was subjected to a tagmentation-based protocol (Nextera XT, Illumina) using one-quarter of the recommended volumes, 10 minuntes for tagmentation at 55 °C and 1 minute extension time during PCR amplification. Libraries were pooled (96 libraries for NextSeq) and sequencing was performed in paired-end mode for 2 × 75 cycles using Illumina's NextSeq 500. Overall design: 47 wild-type samples, 1 wild-type negative-control, 47 H3.V-/- H4.V-/- samples, 1 H3.V-/- H4.V-/- negative control

INSTRUMENT(S): Illumina NextSeq 500 (Trypanosoma brucei brucei)

SUBMITTER: Konrad U. Förstner  

PROVIDER: GSE120515 | GEO | 2018-10-09

REPOSITORIES: GEO

Dataset's files

Source:
Action DRS
GSE120515_Feature_counts_scRNAseq_set_one.txt.gz Txt
GSE120515_VSG_raw_counts_scRNAseq_set_one.txt.gz Txt
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Publications


Many evolutionarily distant pathogenic organisms have evolved similar survival strategies to evade the immune responses of their hosts. These include antigenic variation, through which an infecting organism prevents clearance by periodically altering the identity of proteins that are visible to the immune system of the host<sup>1</sup>. Antigenic variation requires large reservoirs of immunologically diverse antigen genes, which are often generated through homologous recombination, as well as me  ...[more]

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