Project description:Purpose: Evaluation of the m6A modification of PRV and PK15 transcripts during PRV infection Methods: Porcine kidney cell line PK15 was uninfected or infected with PRV for 24 hours. Total RNA from each sample were extracted. Intact mRNA was isolated from total RNA samples and then chemically fragmented to 300-nucleoside-long fragments. Fragmented mRNAs were immunoprecipitated with anti-N6-methyadenosine (m6A) antibody (a part of the fragmented mRNAs was kept as input). Both m6A enriched mRNAs and input mRNAs were concentrated for RNA-seq libraries construction. The libraries were forwarded to sequencing run on Illumina NovaSeq 6000. Results: PRV transcripts were m6A modified during PRV infection and PRV infection changed m6A modification profiles of PK15 transcripts.
Project description:To study in vitro the epithelial cells and PrV interactions during infection, we followed PrV and PK15 cells transcriptome modifications during time-course infection (I) and mock-infection (MI). Four time points were studied: 1h, 2h, 4h and 8h post-I and MI. Four replicates of I and MI were analysed. Keywords: Pig, PrV, Pk15 cells, kinetics
Project description:To study in vitro the epithelial cells and PrV interactions during infection, we followed PrV and PK15 cells transcriptome modifications during time-course infection (I) and mock-infection (MI). Four time points were studied: 1h, 2h, 4h and 8h post-I and MI. Four replicates of I and MI were analysed. 32 samples - The hybridization scheme which can be define as dye-switch was chosen. A balanced loop design with two independent loops, each loop containing 2 replicates of PrV infection and MI, was used. In total, 32 slides were used in this experiment.
Project description:To study in vitro the epithelial cells and PrV interactions during infection, we followed PrV and PK15 cells transcriptome modifications during time-course infection (I) and mock-infection (MI).Six time points were studied: just after I and MI, 1h, 2h, 4h, 8h and 12h post-I and MI. For this study, a pig DNA/cDNA microarray containing genes of the SLA region, additional genes encoding other important immunological molecules and all the PrV genes was constructed. Keywords: infection time course
Project description:Large White and Meishan pigs were either non-treated or injected with mammalian 1-24 ACTH (Immediate Synachten, Novartis France) at the dose of 250 µg per animal. Pigs were sacrificed either immediately after capture from their home cage (non-treated animals) or 1 hour following ACTH injection. Adrenal glands were immediately collected from pigs and frozen on dry ice and then stored at -80°C until RNA isolation. Keywords: stress response, adrenal, gene expression, pig
Project description:BACKGROUND:In animal breeding, identification of causative genetic variants is of major importance and high economical value. Usually, the number of candidate variants exceeds the number of variants that can be validated. One way of prioritizing probable candidates is by evaluating their potential to have a deleterious effect, e.g. by predicting their consequence. Due to experimental difficulties to evaluate variants that do not cause an amino-acid substitution, other prioritization methods are needed. For human genomes, the prediction of deleterious genomic variants has taken a step forward with the introduction of the combined annotation dependent depletion (CADD) method. In theory, this approach can be applied to any species. Here, we present pCADD (p for pig), a model to score single nucleotide variants (SNVs) in pig genomes. RESULTS:To evaluate whether pCADD captures sites with biological meaning, we used transcripts from miRNAs and introns, sequences from genes that are specific for a particular tissue, and the different sites of codons, to test how well pCADD scores differentiate between functional and non-functional elements. Furthermore, we conducted an assessment of examples of non-coding and coding SNVs, which are causal for changes in phenotypes. Our results show that pCADD scores discriminate between functional and non-functional sequences and prioritize functional SNVs, and that pCADD is able to score the different positions in a codon relative to their redundancy. Taken together, these results indicate that based on pCADD scores, regions with biological relevance can be identified and distinguished according to their rate of adaptation. CONCLUSIONS:We present the ability of pCADD to prioritize SNVs in the pig genome with respect to their putative deleteriousness, in accordance to the biological significance of the region in which they are located. We created scores for all possible SNVs, coding and non-coding, for all autosomes and the X chromosome of the pig reference sequence Sscrofa11.1, proposing a toolbox to prioritize variants and evaluate sequences to highlight new sites of interest to explain biological functions that are relevant to animal breeding.
Project description:Large White and Meishan pigs were either non-treated or injected with mammalian 1-24 ACTH (Immediate Synachten, Novartis France) at the dose of 250 µg per animal. Pigs were sacrificed either immediately after capture from their home cage (non-treated animals) or 1 hour following ACTH injection. Adrenal glands were immediately collected from pigs and frozen on dry ice and then stored at -80°C until RNA isolation. Keywords: stress response, adrenal, gene expression, pig 47 samples