Multiple-trait QTL mapping for body and organ weights in a cross between NMRI8 and DB/2 mice

This publication is with permission of the rights owner freely accessible due to an Alliance licence and a national licence (funded by the DFG, German Research Foundation) respectively.Multiple-trait analyses have been shown to improve the detection of quantitative trait loci (QTLs) with multiple effects. Here we applied a multiple-trait approach on obesity- and growth-related traits that were surveyed in 275 F2 mice generated from an intercross between the high body weight selected line NMRI8 and DBA/2 as lean control. The parental lines differed 2·5-fold in body weight at the age of 6 weeks. Within the F2 population, the correlations between body weight and weights of abdominal fat weight, muscle, liver and kidney at the age of 6 weeks were about 0·8. A least squares multiple-trait QTL analysis was performed on these data to understand more precisely the cause of the genetic correlation between body weight, body composition traits and weights of inner organs. Regions on Chr 1, 2, 7 and 14 for body weights at different early ages and regions on Chr 1, 2, 4, 7, 14, 17 and 19 for organ weights at 6 weeks were found to have significant multiple effects at the genome-wide level.Peer Reviewe

Tracking chromosomal positions of oligomers - a case study with Illumina's BovineSNP50 beadchip

<p>Abstract</p> <p>Background</p> <p>High density genotyping arrays have become established as a valuable research tool in human genetics. Currently, more than 300 genome wide association studies were published for human reporting about 1,000 SNPs that are associated with a phenotype. Also in animal sciences high density genotyping arrays are harnessed to analyse genetic variation. To exploit the full potential of this technology single nucleotide polymorphisms (SNPs) on the chips should be well characterized and their chromosomal position should be precisely known. This, however, is a challenge if the genome sequence is still subject to changes.</p> <p>Results</p> <p>We have developed a mapping strategy and a suite of software scripts to update the chromosomal positions of oligomer sequences used for SNP genotyping on high density arrays. We describe the mapping procedure in detail so that scientists with moderate bioinformatics skills can reproduce it. We furthermore present a case study in which we re-mapped 54,001 oligomer sequences from Ilumina's BovineSNP50 beadchip to the bovine genome sequence. We found in 992 cases substantial discrepancies between the manufacturer's annotations and our results. The software scripts in the <monospace>Perl</monospace> and <monospace>R</monospace> programming languages are provided as supplements.</p> <p>Conclusions</p> <p>The positions of oligomer sequences in the genome are volatile even within one build of the genome. To facilitate the analysis of data from a GWAS or from an expression study, especially with species whose genome assembly is still unstable, it is recommended to update the oligomer positions before data analysis.</p

NovelSNPer: A Fast Tool for the Identification and Characterization of Novel SNPs and InDels

Typically, next-generation resequencing projects produce large lists of variants. NovelSNPer is a software tool that permits fast and efficient processing of such output lists. In a first step, NovelSNPer determines if a variant represents a known variant or a previously unknown variant. In a second step, each variant is classified into one of 15 SNP classes or 19 InDel classes. Beside the classes used by Ensembl, we introduce POTENTIAL_START_GAINED and START_LOST as new functional classes and present a classification scheme for InDels. NovelSNPer is based upon the gene structure information stored in Ensembl. It processes two million SNPs in six hours. The tool can be used online or downloaded

BDNF Contributes to the Genetic Variance of Milk Fat Yield in German Holstein Cattle

The gene encoding the brain-derived neurotrophic factor (BDNF) has been repeatedly associated with human obesity. As such, it could also contribute to the regulation of energy partitioning and the amount of secreted milk fat during lactation, which plays an important role in milk production in dairy cattle. Therefore, we performed an association study using estimated breeding values (EBVs) of bulls and yield deviations of German Holstein dairy cattle to test the effect of BDNF on milk fat yield (FY). A highly significant effect (corrected p-value = 3.362 × 10−4) was identified for an SNP 168 kb up-stream of the BDNF transcription start. The association tests provided evidence for an additive allele effect of 5.13 kg of fat per lactation on the EBV for milk FY in bulls and 6.80 kg of fat of the own production performance in cows explaining 1.72 and 0.60% of the phenotypic variance in the analyzed populations, respectively. The analyses of bulls and cows consistently showed three haplotype groups that differed significantly from each other, suggesting at least two different mutations in the BDNF region affecting the milk FY. The FY increasing alleles also had low but significant positive effects on protein and total milk yield which suggests a general role of the BDNF region in energy partitioning, rather than a specific regulation of fat synthesis. The results obtained in dairy cattle suggest similar effects of BDNF on milk composition in other species, including man

Chicken Immune Cell Assay to Model Adaptive Immune Responses In Vitro

Knowledge about the modes of action of immunomodulating compounds such as pathogens, drugs, or feed additives, e.g., probiotics, will allow the development of targeted nutrition strategies, prevent infectious diseases and the usage of antimicrobials, and promote the health of animals. To investigate the mechanisms of action of immunomodulating compounds, controlled in vitro systems using freshly isolated immune cells from blood represent a promising alternative to animal experiments. Immune cell isolation from the blood of chickens is a complex and difficult process since the immune cell fractions are significantly contaminated with red blood cells and platelets. To our knowledge, a robust protocol for immune cell isolation from chicken blood and the subsequent cultivation of immune cells is not available. Here, we established a protocol for blood sampling and immune cell isolation and cultivation from chicken blood, which could be applied for the investigation of direct effects of immunomodulating compounds. This protocol, combining different techniques of immune cell isolation, cultivation, and differentiation of distinct immune cell populations, will serve as a potential alternative to animal testing in vivo. By gaining knowledge about the mechanisms of action of immunomodulating compounds, this in vitro model will contribute to promote health and welfare in chicken farming. Abstract Knowledge about the modes of action of immunomodulating compounds such as pathogens, drugs, or feed additives, e.g., probiotics, gained through controlled but animal-related in vitro systems using primary cultured peripheral blood mononuclear cells (PBMCs) will allow the development of targeted nutrition strategies. Moreover, it could contribute to the prevention of infectious diseases and the usage of antimicrobials, and further promote the health of the animals. However, to our knowledge, a protocol for the isolation of PBMCs with reduced thrombocyte count from chicken blood and subsequent cell culture over several days to assess the effects of immunomodulating compounds is not available. Therefore, we established an optimized protocol for blood sampling and immune cell isolation, culture, and phenotyping for chicken PBMCs. For blood sampling commercial Na–citrate tubes revealed the highest count of vital cells compared to commercial Li–heparin (p < 0.01) and K3EDTA (p < 0.05) tubes. Using combined dextran and ficoll density gradient separation, the thrombocyte count was significantly reduced (p < 0.01) compared to slow-speed centrifugation with subsequent ficoll. For cell culture, the supplementation of RPMI-1640 medium with 10% chicken serum resulted in the lowest relative cell count of thrombocytes compared to fetal calf serum (FCS) (p < 0.05). To validate the ability of the cell culture system to respond to stimuli, concanavalin A (conA) was used as a positive control. The optimized protocol allows the isolation and cultivation of vital PBMCs with reduced thrombocyte count from chicken blood for subsequent investigation of the modes of action of immunomodulating compounds.Peer Reviewe

Finding the Optimal Imputation Strategy for Small Cattle Populations

The imputation from lower density SNP chip genotypes to whole-genome sequence level is an established approach to generate high density genotypes for many individuals. Imputation accuracy is dependent on many factors and for small cattle populations such as the endangered German Black Pied cattle (DSN), determining the optimal imputation strategy is especially challenging since only a low number of high density genotypes is available. In this paper, the accuracy of imputation was explored with regard to (1) phasing of the target population and the reference panel for imputation, (2) comparison of a 1-step imputation approach, where 50 k genotypes are directly imputed to sequence level, to a 2-step imputation approach that used an intermediate step imputing first to 700 k and subsequently to sequence level, (3) the software tools Beagle and Minimac, and (4) the size and composition of the reference panel for imputation. Analyses were performed for 30 DSN and 30 Holstein Frisian cattle available from the 1000 Bull Genomes Project. Imputation accuracy was assessed using a leave-one-out cross validation procedure. We observed that phasing of the target populations and the reference panels affects the imputation accuracy significantly. Minimac reached higher accuracy when imputing using small reference panels, while Beagle performed better with larger reference panels. In contrast to previous research, we found that when a low number of animals is available at the intermediate imputation step, the 1-step imputation approach yielded higher imputation accuracy compared to a 2-step imputation. Overall, the size of the reference panel for imputation is the most important factor leading to higher imputation accuracy, although using a larger reference panel consisting of a related but different breed (Holstein Frisian) significantly reduced imputation accuracy. Our findings provide specific recommendations for populations with a limited number of high density genotyped or sequenced animals available such as DSN. The overall recommendation when imputing a small population are to (1) use a large reference panel of the same breed, (2) use a large reference panel consisting of diverse breeds, or (3) when a large reference panel is not available, we recommend using a smaller same breed reference panel without including a different related breed

Review: Genetic and protein variants of milk caseins in goats

The milk casein genes in goats, are highly polymorphic genes with numerous synonymous and non-synonymous mutations. So far, 20 protein variants have been reported in goats for alpha-S1-casein, eight for beta-casein, 14 for alpha-S2-casein, and 24 for kappa-casein. This review provides a comprehensive overview on identified milk casein protein variants in goat and non-coding DNA sequence variants with some affecting the expression of the casein genes. The high frequency of some casein protein variants in different goat breeds and geographical regions might reflect specific breeding goals with respect to milk processing characteristics, properties for human nutrition and health, or adaptation to the environment. Because protein names, alongside the discovery of protein variants, go through a historical process, we linked old protein names with new ones that reveal more genetic variability. The haplotypes across the cluster of the four genetically linked casein genes are recommended as a valuable genetic tool for discrimination between breeds, managing genetic diversity within and between goat populations, and breeding strategies. The enormous variation in the casein proteins and genes is crucial for producing milk and dairy products with different properties for human health and nutrition, and for genetic improvement depending on local breeding goals

A 5′ UTR Mutation Contributes to Down-Regulation of Bbs7 in the Berlin Fat Mouse

The Bardet–Biedl Syndrome 7 (Bbs7) gene was identified as the most likely candidate gene causing juvenile obesity in the Berlin Fat Mouse Inbred (BFMI) line. Bbs7 expression is significantly lower in the brain, adipose tissue, and liver of BFMI mice compared to lean C57BL/6NCrl (B6N) mice. A DNA sequence comparison between BFMI and B6N revealed 16 sequence variants in the Bbs7 promoter region. Here, we tested if these mutations contribute to the observed differential expression of Bbs7. In a cell-based dual-luciferase assay, we compared the effects of the BFMI and the B6N haplotypes of different regions of the Bbs7 promotor on the reporter gene expression. A single-nucleotide polymorphism (SNP) was identified causing a significant reduction in the reporter gene expression. This SNP (rs29947545) is located in the 5′ UTR of Bbs7 at Chr3:36.613.350. The SNP is not unique to BFMI mice but also occurs in several other mouse strains, where the BFMI allele is not associated with lower Bbs7 transcript amounts. Thus, we suggest a compensatory mutation in the other mouse strains that keeps Bbs7 expression at the normal level. This compensatory mechanism is missing in BFMI mice and the cell lines tested