Gene function & annotation

Identification of coding and splicing-related variants in A1A1 and A2A2 Holstein dairy cows

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Abstract

The A2 β-casein variant has gained considerable interest due to proposed health benefits, contributing to the increasing frequency of the A2A2 genotype in dairy herds. Although the A1/A2 substitution in the β-casein gene (CSN2) does not directly modify gene regulation, previous studies have reported transcriptomic differences, particularly at the mRNA isoform level, between A1A1 and A2A2 cows. Thus, this study aimed to investigate whether Holstein cows with A1A1 and A2A2 β-casein genotypes exhibit distinct variants (SNVs, insertions and deletions) using milk fat globule (MFG) RNA-seq data, and to integrate these results with previously reported mRNA isoform expression data. To minimize potential pedigree-related biases, cows (n = 7 per genotype) were selected as daughters of 12 different sires. Variant calling was performed using CLC Genomics Workbench, and variants were classified according to their predicted effects: (i) amino acid change (AAC), (ii) splice site effect (SSE), and (iii) variants exhibiting both effects. The Ensembl Variant Effect Predictor (VEP) was used to annotate all variants according to their predicted consequence types. Furthermore, only SNVs predicted to induce AAC with SIFT (Sorting Intolerant From Tolerant) scores below 0.05 were retained, as these values indicate a high probability of affecting protein function. Among them, genes identified in the A1A1 group were predominantly associated with mitochondrial oxidative phosphorylation and the electron transport chain (NDUFV2, NDUFAB1, NDUFS8, COX7A2 and ATP5PF), together with genes related to mitochondrial integrity and function (SLC25A44, PISD and IMMT) and fatty-acid activation (ACSL1). Variants identified in A2A2 cows were also located in genes related to mitochondrial integrity and function (OPA1, SLC25A46, VPS13C and SIRT4), lipid metabolism (SPTLC2, LPIN1, FASN, CPT1B and ATP11B) and vesicle trafficking (ABCA2, ABCA5, SEC31A, MVB12B, LRRK2). Moreover, variants predicted as SSE were integrated with previously reported DE mRNA isoforms, and only those classified as having High or Moderate impact by VEP were further analysed. A total of 85 SNVs, 65 insertions and 74 deletions were found, highlighting the interest of integrate analysis of mRNA isoforms and RNA-seq variant calling. A deeper in vivo functional validation of these variants, identified in expressed regions of the genome, may help to clarify their biological relevance in the context of selective breeding for the A2A2 β-casein genotype in Holstein dairy cows.

Keywords: 2026

How to Cite:

Cánovas, à., Jiménez Montenegro, L. & Urrutia Vera, O., (2026) “Identification of coding and splicing-related variants in A1A1 and A2A2 Holstein dairy cows”, World Congress on Genetics Applied to Livestock Production Digital Archive 2026(1): 2287348. doi: https://doi.org/10.31274/wcgalp.24273

Rights: 1