A frameshift mutation in ARMC3 is associated with a tail stump sperm defect in Swedish Red (Bos taurus) cattle.

BACKGROUND: Artificial insemination is widely used in many cattle breeding programs. Semen samples of breeding bulls are collected and closely examined immediately after collection at artificial insemination centers. Only ejaculates without anomalous findings are retained for artificial insemination. Although morphological aberrations of the spermatozoa are a frequent reason for discarding ejaculates, the genetic determinants underlying poor semen quality are scarcely understood. RESULTS: A tail stump sperm defect was observed in three bulls of the Swedish Red cattle breed. The spermatozoa of affected bulls were immotile because of severely disorganized tails indicating disturbed spermatogenesis. We genotyped three affected bulls and 18 unaffected male half-sibs at 46,035 SNPs and performed homozygosity mapping to map the fertility disorder to an 8.42 Mb interval on bovine chromosome 13. The analysis of whole-genome re-sequencing data of an affected bull and 300 unaffected animals from eleven cattle breeds other than Swedish Red revealed a 1 bp deletion (Chr13: 24,301,425 bp, ss1815612719) in the eleventh exon of the armadillo repeat containing 3-encoding gene (ARMC3) that was compatible with the supposed recessive mode of inheritance. The deletion is expected to alter the reading frame and to induce premature translation termination (p.A451fs26). The mutated protein is shortened by 401 amino acids (46 %) and lacks domains that are likely essential for normal protein function. CONCLUSIONS: We report the phenotypic and genetic characterization of a sterilizing tail stump sperm defect in the Swedish Red cattle breed. Exploiting high-density genotypes and massive re-sequencing data enabled us to identify the most likely causal mutation for the fertility disorder in bovine ARMC3. Our results provide the basis for monitoring the mutated variant in the Swedish Red cattle population and for the early identification of infertile animals.

A robust linkage map of the porcine autosomes based on gene-associated SNPs.

BACKGROUND: Genetic linkage maps are necessary for mapping of mendelian traits and quantitative trait loci (QTLs). To identify the actual genes, which control these traits, a map based on gene-associated single nucleotide polymorphism (SNP) markers is highly valuable. In this study, the SNPs were genotyped in a large family material comprising more than 5,000 piglets derived from 12 Duroc boars crossed with 236 Danish Landrace/Danish Large White sows. The SNPs were identified in sequence alignments of 4,600 different amplicons obtained from the 12 boars and containing coding regions of genes derived from expressed sequence tags (ESTs) and genomic shotgun sequences. RESULTS: Linkage maps of all 18 porcine autosomes were constructed based on 456 gene-associated and six porcine EST-based SNPs. The total length of the averaged-sex whole porcine autosome was estimated to 1,711.8 cM resulting in an average SNP spacing of 3.94 cM. The female and male maps were estimated to 2,336.1 and 1,441.5 cM, respectively. The gene order was validated through comparisons to the cytogenetic and/or physical location of 203 genes, linkage to evenly spaced microsatellite markers as well as previously reported conserved synteny. A total of 330 previously unmapped genes and ESTs were mapped to the porcine autosome while ten genes were mapped to unexpected locations. CONCLUSION: The linkage map presented here shows high accuracy in gene order. The pedigree family network as well as the large amount of meiotic events provide good reliability and make this map suitable for QTL and association studies. In addition, the linkage to the RH-map of microsatellites makes it suitable for comparison to other QTL studies.

Gene expression profiling, chromosome assignment and mutational analysis of the porcine Golgi-resident UDP-N-acetylglucosamine transporter SLC35A3.

SLC35A3 encodes a Golgi-resident UDP-N-acetylglucosamine transporter. Here, the porcine SLC35A3 gene was assigned to Sus scrofa chromosome 4 (SSC4) by a combination of radiation hybrid and linkage analysis. Expression profiling using real time RT-PCR showed ubiquitous but variable transcription of SLC35A3 in a selection of tissues. The deduced 325 amino acid sequence revealed a hydrophobic protein with 10 predicted transmembrane helices and the N- and C-terminal tails facing the cytosolic side of the Golgi apparatus. In addition, mutated versions of the UDP-GlcNAc transporter were analyzed in a yeast complementation assay, which allowed us to identify important domains and amino acid residues. Thus, the N-terminal tail was inessential for activity, whereas removal of the first transmembrane domain inhibited yeast complementation. The hydrophilic C-terminus was dispensable while mutant proteins either fully or partially deprived of the last membrane-spanning helix were functionally impaired. The third luminal loop showed modest sequence conservation and appeared structurally flexible as certain deletions were acceptable. In contrast, the fourth luminal loop was more sensitive to changes since the competence of the mutant protein was lowered by mutations. Substitutions of glycines 190, 215 and 254, which are invariant positions in the SLC35A subfamilies affected activity negatively. Interestingly, inhibition of function by a valine to phenylalanine mutation, which has been associated with skeletal malformations, is likely caused by structural incompatibility of the bulky aromatic phenylalanine side chain with the integrity of the transmembrane helix, since substitutions with the smaller aliphatic side chains of leucine and isoleucine were acceptable changes.

SNP mining porcine ESTs with MAVIANT, a novel tool for SNP evaluation and annotation.

MOTIVATION: Single nucleotide polymorphisms (SNPs) analysis is an important means to study genetic variation. A fast and cost-efficient approach to identify large numbers of novel candidates is the SNP mining of large scale sequencing projects. The increasing availability of sequence trace data in public repositories makes it feasible to evaluate SNP predictions on the DNA chromatogram level. MAVIANT, a platform-independent Multipurpose Alignment VIewing and Annotation Tool, provides DNA chromatogram and alignment views and facilitates evaluation of predictions. In addition, it supports direct manual annotation, which is immediately accessible and can be easily shared with external collaborators. RESULTS: Large-scale SNP mining of polymorphisms bases on porcine EST sequences yielded more than 7900 candidate SNPs in coding regions (cSNPs), which were annotated relative to the human genome. Non-synonymous SNPs were analyzed for their potential effect on the protein structure/function using the PolyPhen and SIFT prediction programs. Predicted SNPs and annotations are stored in a web-based database. Using MAVIANT SNPs can visually be verified based on the DNA sequencing traces. A subset of candidate SNPs was selected for experimental validation by resequencing and genotyping. This study provides a web-based DNA chromatogram and contig browser that facilitates the evaluation and selection of candidate SNPs, which can be applied as genetic markers for genome wide genetic studies. AVAILABILITY: The stand-alone version of MAVIANT program for local use is freely available under GPL license terms at http://snp.agrsci.dk/maviant. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.