Allele specific expression in various tissues of Gallus gallus domesticus
Date
2018
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Publisher
University of Delaware
Abstract
Allele specific expression (ASE) is the process where one allele in a heterozygous individual is expressed at a higher level in comparison to the other when equal expression is expected. ASE is of great interest because it helps in the identification of cis-regulatory mutations or epigenetic modifications that influence gene expression. In this study, the effect of these cis-regulatory elements is investigated by examining single nucleotide polymorphisms (SNPs) from RNA-seq. Identified SNPs may then be utilized in future animal breeding programs. In our study, we performed RNA-sequencing on 100 samples collected from various populations of chickens. We then followed Genome Analysis Toolkit’s (GATK) “Best Practices for Variant Calling on RNA-seq” using recommended settings. We aligned the sequence reads to the chicken reference genome sequence (Gallus gallus 5.0) from Ensembl. Based on the first round of alignment of STAR, the average number of reads was 36,196,012 with an average mapping rate of 85%. We identified a total 3,147,284 variants (SNPs and Indels) and then used these variants to mask the reference genome for initial alignment and re-ran the pipeline. The final variants from a large sub-set of the prior samples (n= 68), which consisted of samples from different tissues (breast muscle, abdominal fat, liver), but from the same population were examined for ASE. ASE analysis was performed using the custom analysis software called VCF ASE Detection Tool (VADT). It should be mentioned that VCF refers to a variant call file. VADT automatically filters the variant data based on user parameters, detects ASE using a binomial test and automatically performs statistical correction of the results. On average ~174,000 SNPs in each tissue passed our filtering criteria and were considered informative, of which ~24,000 (~14%) showing ASE. The overlap of ASE SNPs among the 3 tissues was only 3.7%, with ~83% of variants showing tissue specificity. Now if variants are mapped to genes and the genes are compared between the tissues, this overlap increases to 20.1%. Overall it was found that ASE genes show enrichment for tissue specificity but were also found to show enrichment for pathways involved with translation and ribosomes. When the top ASE genes that were found in all three tissues were examined, there was a clear enrichment for KEGG pathways involved in ribosomes and metabolism