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Bioinformatic pipelines are becoming increasingly complex with the ever-accumulating amount of Next-generation sequencing (NGS) data. Their orchestration is difficult with a simple Bash script, but bioinformatics workflow managers such as Nextflow provide a framework to overcome respective problems. This study used Nextflow to develop a bioinformatic pipeline for detecting expression quantitative trait loci (eQTL) using a DSL2 Nextflow modular syntax, to enable sharing the huge demand for computing power as well as data access limitation across different partners often associated with eQTL studies. Based on the results from a test run with pilot data by measuring the required runtime and computational resources, the new pipeline should be suitable for eQTL studies in large scale analyses., (© The Author(s) 2024. Published by Oxford University Press on behalf of NAR Genomics and Bioinformatics.)

Understanding the genomic control of tissue-specific gene expression and regulation can help to inform the application of genomic technologies in farm animal breeding programs. The fine mapping of promoters [transcription start sites (TSS)] and enhancers (divergent amplifying segments of the genome local to TSS) in different populations of cattle across a wide diversity of tissues provides information to locate and understand the genomic drivers of breed- and tissue-specific characteristics. To this aim, we used Cap Analysis Gene Expression (CAGE) sequencing, of 24 different tissues from 3 populations of cattle, to define TSS and their coexpressed short-range enhancers (<1 kb) in the ARS-UCD1.2&#95;Btau5.0.1Y reference genome (1000bulls run9) and analyzed tissue and population specificity of expressed promoters. We identified 51,295 TSS and 2,328 TSS-Enhancer regions shared across the 3 populations (dairy, beef-dairy cross, and Canadian Kinsella composite cattle from 2 individuals, 1 of each sex, per population). Cross-species comparative analysis of CAGE data from 7 other species, including sheep, revealed a set of TSS and TSS-Enhancers that were specific to cattle. The CAGE data set will be combined with other transcriptomic information for the same tissues to create a new high-resolution map of transcript diversity across tissues and populations in cattle for the BovReg project. Here we provide the CAGE data set and annotation tracks for TSS and TSS-Enhancers in the cattle genome. This new annotation information will improve our understanding of the drivers of gene expression and regulation in cattle and help to inform the application of genomic technologies in breeding programs., Competing Interests: Conflicts of interest No commercial or academic conflict of interest were declared by any of the authors for this manuscript., (© The Author(s) 2023. Published by Oxford University Press on behalf of The Genetics Society of America.)

Population-wide associations between loci due to linkage disequilibrium can be used to map quantitative trait loci (QTL) with high resolution. However, spurious associations between markers and QTL can also arise as a consequence of population stratification. Statistical methods that cannot differentiate between loci associations due to linkage disequilibria from those caused in other ways can render false-positive results. The transmission-disequilibrium test (TDT) is a robust test for detecting QTL. The TDT exploits within-family associations that are not affected by population stratification. However, some TDTs are formulated in a rigid form, with reduced potential applications. In this study we generalize TDT using mixed linear models to allow greater statistical flexibility. Allelic effects are estimated with two independent parameters: one exploiting the robust within-family information and the other the potentially biased between-family information. A significant difference between these two parameters can be used as evidence for spurious association. This methodology was then used to test the effects of the fourth melanocortin receptor (MC4R) on production traits in the pig. The new analyses supported the previously reported results; i.e., the studied polymorphism is either causal or in very strong linkage disequilibrium with the causal mutation, and provided no evidence for spurious association.

Several quantitative trait loci (QTL) affecting muscle glycogen content and related traits were mapped to pig chromosome 15 using a three-generation intercross between Berkshire x Yorkshire pigs. On the basis of the QTL location the PRKAG3 (protein kinase, AMP-activated, gamma(3)-subunit) gene was considered to be a good candidate for the observed effects. Differences in the PRKAG3 gene sequences of the founder animals of the intercross were analyzed. The RN(-) mutation previously reported was not present in the cross but three missense substitutions and a polymorphic short interspersed element (SINE) were identified. To confirm the hypothesis that at least one of these mutations was associated with differences in meat quality, >1800 animals from several unrelated commercial lines were genotyped for the candidate substitutions and an association study was performed. The results demonstrate the presence of new economically important alleles of the PRKAG3 gene affecting the glycogen content in the muscle and the resulting meat quality. Haplotype analysis was shown to resolve the effects of PRKAG3 more clearly than analysis of individual polymorphisms. Because of their prevalence in the more common commercial breeds, the potential implications for the pig industry and consumers are considerably greater than the original discovery of the RN(-) mutation. Furthermore, these results illustrate that additional alleles of genes involved in major mutations may play a significant role in quantitative trait variation.

Black spotting on a red or white background in pigs is determined by the E(P) allele at the MC1R/Extension locus. A previous comparison of partial MC1R sequences revealed that E(P) shares a missense mutation (D121N) with the E(D2) allele for dominant black color. Sequence analysis of the entire coding region now reveals a second mutation in the form of a 2-bp insertion at codon 23 (nt67insCC). This mutation expands a tract of six C nucleotides to eight and introduces a premature stop codon at position 56. This frameshift mutation is expected to cause a recessive red color, which was in fact observed in some breeds with the E(P) allele present (Tamworth and Hereford). RT-PCR analyses were conducted using skin samples taken from both spotted and background areas of spotted pigs. The background red area had transcript only from the mutant nt67insCC MC1R allele, whereas the black spot also contained a transcript without the 2-bp insertion. This indicates that black spots are due to somatic reversion events that restore the frame and MC1R function. The phenotypic expression of the E(P) allele is highly variable and the associated coat color ranges from red, red with black spots, white with black spots, to almost completely solid black. In several breeds of pigs the phenotypic manifestation of this allele has been modified by selection for or against black spots.