GSE100203
RNA sequencing analysis of the Lewis versus Brown Norway rat, with or without Toxoplasma gondii infection
Rats vary in their susceptibilities to Toxoplasma gondii infection depending on the rat strain. Compared to the T. gondii-susceptible Brown Norway (BN) rat, the Lewis (LEW) rat is extremely resistant to T. gondii. Thus, these two rat strains are ideal models for elucidating host mechanisms that are important for host resistance to T. gondii infection. Therefore, in an attempt to unravel molecular factors directing the protective early innate immune responses in the LEW rat, we performed RNA sequencing analysis of the LEW versus BN rat, with or without T. gondii infection. Overall design: Total RNA was obtained from fresh peritoneal cells of LEW and BN rats (with or without T. gondii infection), 24 hours post-infection. RNA libraries were generated using the Illumina''s ''TruSeq Stranded mRNAseq Sample Prep kit'' (Illumina). The libraries were quantitated by fluorometry (Qubit), and their quality determined using a Bioanalyzer. The libraries were then diluted and quantified by qPCR. Sequencing of the RNA libraries was done using a HiSeq SBS sequencing kit version 4 and a HiSeq2500 system (Illumina). Residual adapter content and low quality bases on the generated sequences were trimmed using Trimmomatic (version 0.33), while the bcl2fastq v2.17.1.14 Conversion Software (Illumina) was used to generate and demultiplex Fastq files from the sequence data. ASCII offset of 33 known as Sanger scores were used to determine the quality scores line in fastq files. Feature Counts in the Sub-read (version 1.5.0) package were used to derive gene counts, while gene annotations, including gene ontology terms, were downloaded from Ensembl Release 84 for Rnor_6.0, and ToxoDB.org Release 28 for T. gondii GT1. Samtools (v. 1.3) idxstats tool was used to determine the number of reads mapping to each rat chromosome and to the T. gondii genome. By ANOVA, using a false discovery rate (FDR) of 0.05, genes' expression patterns related to the four experimental groups (T. gondii-infected and uninfected LEW and BN rat groups; n = 4 per group) were generated with reads ranging from 13.1 – 18.3 million per sample that aligned uniquely within 32,662 rat genes and 8,637 T. gondii genes.