Differential Genes Expression of Blood Tissue Related to Pre-Calving Ketosis in Holstein cow using Transcriptomics Data

In recent decades, dairy cows have been selected for high levels of milk production; As a result, the cow's demand for nutrients increases so much that if the required amount of energy is not provided, the incidence of ketosis in dairy cows will increase; Therefore, in this study, the aim is to identify different index genes expressed between healthy and pre-calving ketosis groups that ultimately identify the metabolic pathways and molecular activities involved during ketosis.

Transcript (total mRNA sequence) of six blood tissue samples containing three replicates from each group was prepared. The preprocessing steps of the readings were performed using Fastq-dump, FastQC and Trimmomatic software and also the study of gene expression profile was performed using Hisat2 and Samtools software. Finally, the differential expression analysis of genes was performed using Cufflinks software package and gene ontology analysis was performed using DAVID and Ensembl databases.

Finally, 35342 genes and 107722 isoforms were identified on the transcript of these samples and finally 179 genes with different and significant expression were identified (p <0.00055) which, considering log2> 4 and log2 <-4, Seven genes with different expression between the two groups were studied. The largest differences in gene expression between the two groups were related to the genes, HBA and PTGS2 with log2 (fold_change), 4.53 and 4.47, respectively. Gene ontology (GO) analysis and the pathways involved in it showed that the mitochondrial membrane and endoplasmic reticulum component, inflammatory response and oxidative stress, hemoglobin redox process and biosynthesis, cellular response to hypoxia, metal ion binding, biosynthesis and metabolism Fatty acids and lipids are also involved in the chemokine and TNF signaling pathways.

Seven index genes with different and significant expression were identified on the transcript of the samples; The CXCL3 and GRO1 genes had the highest expression between the two groups and the HBA gene had the highest differential expression, with more than 23-fold expression in the pre-calving ketosis group compared to the healthy group. Gene ontology analysis showed that HBA gene as the most significant gene with different expression, with more expression in the group with pre-calving ketosis, is a factor in increasing oxygen transfer, hemoglobin biosynthesis, oxygen binding and iron ions.