Publications by authors named "Rudiger Brauning"

30 Publications

  • Page 1 of 1

Genomic signatures of inbreeding in a critically endangered parrot, the kākāpō.

G3 (Bethesda) 2021 Aug 31. Epub 2021 Aug 31.

Department of Zoology, University of Otago, Dunedin 9054, New Zealand.

Events of inbreeding are inevitable in critically endangered species. Reduced population sizes and unique life-history traits can increase the severity of inbreeding, leading to declines in fitness and increased risk of extinction. Here, we investigate levels of inbreeding in a critically endangered flightless parrot, the kākāpō (Strigops habroptilus), wherein a highly inbred island population and one individual from the mainland of New Zealand founded the entire extant population. Genotyping-by-sequencing (GBS), and a genotype calling approach using a chromosome-level genome assembly, identified a filtered set of 12,241 single-nucleotide polymorphisms (SNPs) among 161 kākāpō, which together encompass the total genetic potential of the extant population. Multiple molecular-based estimates of inbreeding were compared, including genome-wide estimates of heterozygosity (FH), the diagonal elements of a genomic-relatedness matrix (FGRM), and runs of homozygosity (RoH, FRoH). In addition, we compared levels of inbreeding in chicks from a recent breeding season to examine if inbreeding is associated with offspring survival. The density of SNPs generated with GBS was sufficient to identify chromosomes that were largely homozygous with RoH distributed in similar patterns to other inbred species. Measures of inbreeding were largely correlated and differed significantly between descendants of the two founding populations. However, neither inbreeding nor ancestry was found to be associated with reduced survivorship in chicks, owing to unexpected mortality in chicks exhibiting low levels of inbreeding. Our study highlights important considerations for estimating inbreeding in critically endangered species, such as the impacts of small population sizes and admixture between diverse lineages.
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http://dx.doi.org/10.1093/g3journal/jkab307DOI Listing
August 2021

A Multi-omic Huntington's Disease Transgenic Sheep-Model Database for Investigating Disease Pathogenesis.

J Huntingtons Dis 2021 Aug 18. Epub 2021 Aug 18.

Centre for Brain Research, School of Biological Sciences, The University of Auckland, Auckland, New Zealand.

Background: The pathological mechanism of cellular dysfunction and death in Huntington's disease (HD) is not well defined. Our transgenic HD sheep model (OVT73) was generated to investigate these mechanisms and for therapeutic testing. One particular cohort of animals has undergone focused investigation resulting in a large interrelated multi-omic dataset, with statistically significant changes observed comparing OVT73 and control 'omic' profiles and reported in literature.

Objective: Here we make this dataset publicly available for the advancement of HD pathogenic mechanism discovery.

Methods: To enable investigation in a user-friendly format, we integrated seven multi-omic datasets from a cohort of 5-year-old OVT73 (n = 6) and control (n = 6) sheep into a single database utilising the programming language R. It includes high-throughput transcriptomic, metabolomic and proteomic data from blood, brain, and other tissues.

Results: We present the 'multi-omic' HD sheep database as a queriable web-based platform that can be used by the wider HD research community (https://hdsheep.cer.auckland.ac.nz/). The database is supported with a suite of simple automated statistical analysis functions for rapid exploratory analyses. We present examples of its use that validates the integrity relative to results previously reported. The data may also be downloaded for user determined analysis.

Conclusion: We propose the use of this online database as a hypothesis generator and method to confirm/refute findings made from patient samples and alternate model systems, to expand our understanding of HD pathogenesis. Importantly, additional tissue samples are available for further investigation of this cohort.
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http://dx.doi.org/10.3233/JHD-210482DOI Listing
August 2021

Application of Low Coverage Genotyping by Sequencing in Selectively Bred Arctic Charr ().

G3 (Bethesda) 2020 06 1;10(6):2069-2078. Epub 2020 Jun 1.

Department of Animal Breeding and Genetics, Swedish University of Agricultural Sciences, Box 7090, 750 07 Uppsala, Sweden.

Arctic charr () is a species of high economic value for the aquaculture industry, and of high ecological value due to its Holarctic distribution in both marine and freshwater environments. Novel genome sequencing approaches enable the study of population and quantitative genetic parameters even on species with limited or no prior genomic resources. Low coverage genotyping by sequencing (GBS) was applied in a selected strain of Arctic charr in Sweden originating from a landlocked freshwater population. For the needs of the current study, animals from year classes 2013 (171 animals, parental population) and 2017 (759 animals; 13 full sib families) were used as a template for identifying genome wide single nucleotide polymorphisms (SNPs). GBS libraries were constructed using the PstI and MspI restriction enzymes. Approximately 14.5K SNPs passed quality control and were used for estimating a genomic relationship matrix. Thereafter a wide range of analyses were conducted in order to gain insights regarding genetic diversity and investigate the efficiency of the genomic information for parentage assignment and breeding value estimation. Heterozygosity estimates for both year classes suggested a slight excess of heterozygotes. Furthermore, F estimates among the families of year class 2017 ranged between 0.009 - 0.066. Principal components analysis (PCA) and discriminant analysis of principal components (DAPC) were applied aiming to identify the existence of genetic clusters among the studied population. Results obtained were in accordance with pedigree records allowing the identification of individual families. Additionally, DNA parentage verification was performed, with results in accordance with the pedigree records with the exception of a putative dam where full sib genotypes suggested a potential recording error. Breeding value estimation for juvenile growth through the usage of the estimated genomic relationship matrix clearly outperformed the pedigree equivalent in terms of prediction accuracy (0.51 opposed to 0.31). Overall, low coverage GBS has proven to be a cost-effective genotyping platform that is expected to boost the selection efficiency of the Arctic charr breeding program.
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http://dx.doi.org/10.1534/g3.120.401295DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7263669PMC
June 2020

A restriction enzyme reduced representation sequencing approach for low-cost, high-throughput metagenome profiling.

PLoS One 2020 3;15(4):e0219882. Epub 2020 Apr 3.

AgResearch Limited, Invermay Agricultural Centre, Mosgiel, New Zealand.

Microbial community profiles have been associated with a variety of traits, including methane emissions in livestock. These profiles can be difficult and expensive to obtain for thousands of samples (e.g. for accurate association of microbial profiles with traits), therefore the objective of this work was to develop a low-cost, high-throughput approach to capture the diversity of the rumen microbiome. Restriction enzyme reduced representation sequencing (RE-RRS) using ApeKI or PstI, and two bioinformatic pipelines (reference-based and reference-free) were compared to bacterial 16S rRNA gene sequencing using repeated samples collected two weeks apart from 118 sheep that were phenotypically extreme (60 high and 58 low) for methane emitted per kg dry matter intake (n = 236). DNA was extracted from freeze-dried rumen samples using a phenol chloroform and bead-beating protocol prior to RE-RRS. The resulting sequences were used to investigate the repeatability of the rumen microbial community profiles, the effect of laboratory and analytical method, and the relationship with methane production. The results suggested that the best method was PstI RE-RRS analyzed with the reference-free approach, which accounted for 53.3±5.9% of reads, and had repeatabilities of 0.49±0.07 and 0.50±0.07 for the first two principal components (PC1 and PC2), phenotypic correlations with methane yield of 0.43±0.06 and 0.46±0.06 for PC1 and PC2, and explained 41±8% of the variation in methane yield. These results were significantly better than for bacterial 16S rRNA gene sequencing of the same samples (p<0.05) except for the correlation between PC2 and methane yield. A Sensitivity study suggested approximately 2000 samples could be sequenced in a single lane on an Illumina HiSeq 2500, meaning the current work using 118 samples/lane and future proposed 384 samples/lane are well within that threshold. With minor adaptations, our approach could be used to obtain microbial profiles from other metagenomic samples.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0219882PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7122713PMC
July 2020

GBS Data Identify Pigmentation-Specific Genes of Potential Role in Skin-Photosensitization in Two Tunisian Sheep Breeds.

Animals (Basel) 2019 Dec 18;10(1). Epub 2019 Dec 18.

National Agricultural Research Institute of Tunisia, Laboratory of Animal and forage Production, University of Carthage, Ariana 1004, Tunisia.

The Tunisian Noire de Thibar sheep breed is a composite breed, recently selected to create animals that are uniformly black in order to avoid skin photosensitization after the ingestion of toxic "" weeds, which causes a major economic loss to sheep farmers. We assessed genetic differentiation and estimated marker F using genotyping-by-sequencing (GBS) data in black (Noire de Thibar) and related white-coated (Queue fine de l'ouest) sheep breeds to identify signals of artificial selection. The results revealed the selection signatures within candidate genes related to coat color, which are assumed to be indirectly involved in the mechanism of photosensitization in sheep. The identified genes could provide important information for molecular breeding.
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http://dx.doi.org/10.3390/ani10010005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7022847PMC
December 2019

Exclusion and Genomic Relatedness Methods for Assignment of Parentage Using Genotyping-by-Sequencing Data.

G3 (Bethesda) 2019 10 7;9(10):3239-3247. Epub 2019 Oct 7.

AgResearch, Invermay Agricultural Centre, Private Bag 50034, Mosgiel 9053, New Zealand.

Genotypes are often used to assign parentage in agricultural and ecological settings. Sequencing can be used to obtain genotypes but does not provide unambiguous genotype calls, especially when sequencing depth is low in order to reduce costs. In that case, standard parentage analysis methods no longer apply. A strategy for using low-depth sequencing data for parentage assignment is developed here. It entails the use of relatedness estimates along with a metric termed excess mismatch rate which, for parent-offspring pairs or trios, is the difference between the observed mismatch rate and the rate expected under a model of inheritance and allele reads without error. When more than one putative parent has similar statistics, bootstrapping can provide a measure of the relatedness similarity. Putative parent-offspring trios can be further checked for consistency by comparing the offspring's estimated inbreeding to half the parent relatedness. Suitable thresholds are required for each metric. These methods were applied to a deer breeding operation consisting of two herds of different breeds. Relatedness estimates were more in line with expectation when the herds were analyzed separately than when combined, although this did not alter which parents were the best matches with each offspring. Parentage results were largely consistent with those based on a microsatellite parentage panel with three discordant parent assignments out of 1561. Two models are investigated to allow the parentage metrics to be calculated with non-random selection of alleles. The tools and strategies given here allow parentage to be assigned from low-depth sequencing data.
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http://dx.doi.org/10.1534/g3.119.400501DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6778805PMC
October 2019

Complete Genome Sequence of the Telford Type S Strain of Mycobacterium avium subsp. paratuberculosis.

Microbiol Resour Announc 2019 Mar 14;8(11). Epub 2019 Mar 14.

AgResearch Ltd., Hopkirk Research Institute, Palmerston North, New Zealand

subsp. is the causative agent of Johne's disease (JD). Here, we report the complete genome sequence of Telford 9.2, a well-characterized representative strain of the subsp. S subtype that is endemic in New Zealand and Australian sheep.
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http://dx.doi.org/10.1128/MRA.00004-19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6424202PMC
March 2019

Whole Genome Sequencing for Determining the Source of Infections in Livestock Herds and Wildlife in New Zealand.

Front Vet Sci 2018 30;5:272. Epub 2018 Oct 30.

AgResearch, Hopkirk Research Institute, Palmerston North, New Zealand.

The ability to DNA fingerprint isolates helped to define the role of wildlife in the persistence of bovine tuberculosis in New Zealand. DNA fingerprinting results currently help to guide wildlife control measures and also aid in tracing the source of infections that result from movement of livestock. During the last 5 years we have developed the ability to distinguish New Zealand (NZ) isolates by comparing the sequences of whole genome sequenced (WGS) samples. WGS provides much higher resolution than our other established typing methods and greatly improves the definition of the regional localization of NZ types. Three outbreak investigations are described and results demonstrate how WGS analysis has led to the confirmation of epidemiological sourcing of infection, to better definition of new sources of infection by ruling out other possible sources, and has revealed probable wildlife infection in an area considered to be free of infected wildlife. The routine use of WGS analyses for sourcing new infections will be an important component of the strategy employed to eradicate bovine TB from NZ livestock and wildlife.
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http://dx.doi.org/10.3389/fvets.2018.00272DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6218598PMC
October 2018

Population Connectivity and Traces of Mitochondrial Introgression in New Zealand Black-Billed Gulls ().

Genes (Basel) 2018 Nov 9;9(11). Epub 2018 Nov 9.

Department of Zoology, University of Otago, Great King Street, Dunedin 9016, New Zealand.

Black-billed gulls () are endemic to New Zealand and are suspected to be undergoing substantial population declines. They primarily breed on open gravel beds in braided rivers of the South Island-a habitat that is diminishing and becoming increasingly modified. Although management of this species is increasing, little has been published on their movements and demographics. In this study, both mitochondrial DNA (mtDNA) control region domain I and nuclear single nucleotide polymorphisms (SNPs) were examined to help understand the connectivity and population structure of black-billed gulls across the country and to help inform management decisions. Mitochondrial DNA showed no population structure, with high haplotype and low nucleotide diversity, and analyses highlighted mitochondrial introgression with the closely related red-billed gulls (). Nuclear DNA analyses, however, identified two groups, with Rotorua birds in the North Island being distinct from the rest of New Zealand, and isolation-by-distance evident across the South Island populations. Gene flow primarily occurs between nearby colonies with a stepwise movement across the landscape. The importance from a genetic perspective of the more isolated North Island birds (1.6% of total population) needs to be further evaluated. From our results, we infer that the South Island black-billed gull management should focus on maintaining several populations within each region rather than focusing on single specific colonies or river catchments. Future study is needed to investigate the genetic structure of populations at the northern limit of the species' range, and identify the mechanisms behind, and extent of, the hybridisation between red-billed and black-billed gulls.
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http://dx.doi.org/10.3390/genes9110544DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6266082PMC
November 2018

An entropy-reducing data representation approach for bioinformatic data.

Database (Oxford) 2018 01;2018

AgResearch, Invermay Agricultural Centre, Mosgiel, New Zealand.

Database Url: https://github.com/AgResearch/data_prism.
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http://dx.doi.org/10.1093/database/bay029DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5887302PMC
January 2018

Linkage Disequilibrium Estimation in Low Coverage High-Throughput Sequencing Data.

Genetics 2018 06 27;209(2):389-400. Epub 2018 Mar 27.

AgResearch, Invermay Agricultural Centre, Mosgiel 9053, New Zealand.

High-throughput sequencing methods that multiplex a large number of individuals have provided a cost-effective approach for discovering genome-wide genetic variation in large populations. These sequencing methods are increasingly being utilized in population genetic studies across a diverse range of species. Two side-effects of these methods, however, are (1) sequencing errors and (2) heterozygous genotypes called as homozygous due to only one allele at a particular locus being sequenced, which occurs when the sequencing depth is insufficient. Both of these errors have a profound effect on the estimation of linkage disequilibrium (LD) and, if not taken into account, lead to inaccurate estimates. We developed a new likelihood method, GUS-LD, to estimate pairwise linkage disequilibrium using low coverage sequencing data that accounts for undercalled heterozygous genotypes and sequencing errors. Our findings show that accurate estimates were obtained using GUS-LD, whereas underestimation of LD results if no adjustment is made for the errors.
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http://dx.doi.org/10.1534/genetics.118.300831DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5972415PMC
June 2018

Sheep genome functional annotation reveals proximal regulatory elements contributed to the evolution of modern breeds.

Nat Commun 2018 02 28;9(1):859. Epub 2018 Feb 28.

CSIRO Agriculture and Food, 306 Carmody Road, St. Lucia, 4067, QLD, Australia.

Domestication fundamentally reshaped animal morphology, physiology and behaviour, offering the opportunity to investigate the molecular processes driving evolutionary change. Here we assess sheep domestication and artificial selection by comparing genome sequence from 43 modern breeds (Ovis aries) and their Asian mouflon ancestor (O. orientalis) to identify selection sweeps. Next, we provide a comparative functional annotation of the sheep genome, validated using experimental ChIP-Seq of sheep tissue. Using these annotations, we evaluate the impact of selection and domestication on regulatory sequences and find that sweeps are significantly enriched for protein coding genes, proximal regulatory elements of genes and genome features associated with active transcription. Finally, we find individual sites displaying strong allele frequency divergence are enriched for the same regulatory features. Our data demonstrate that remodelling of gene expression is likely to have been one of the evolutionary forces that drove phenotypic diversification of this common livestock species.
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http://dx.doi.org/10.1038/s41467-017-02809-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5830443PMC
February 2018

The effects of transcription and recombination on mutational dynamics of short tandem repeats.

Nucleic Acids Res 2018 02;46(3):1321-1330

Department of Anatomy, University of Otago, Dunedin 9054, New Zealand.

Short tandem repeats (STR) are ubiquitous components of the genomic architecture of most living organisms. Recent work has highlighted the widespread functional significance of such repeats, particularly around gene regulation, but the mutational processes underlying the evolution of these highly abundant and highly variable sequences are not fully understood. Traditional models assume that strand misalignment during replication is the predominant mechanism, but empirical data suggest the involvement of other processes including recombination and transcription. Despite this evidence, the relative influences of these processes have not previously been tested experimentally on a genome-wide scale. Using deep sequencing, we identify mutations at >200 microsatellites, across 700 generations in replicated populations of two otherwise identical sexual and asexual Saccharomyces cerevisiae strains. Using generalized linear models, we investigate correlates of STR mutability including the nature of the mutation, STR composition and contextual factors including recombination, transcription and replication origins. Sexual capability was not a significant predictor of microsatellite mutability, but, intriguingly, we identify transcription as a significant positive predictor. We also find that STR density is substantially increased in regions neighboring, but not within, recombination hotspots.
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http://dx.doi.org/10.1093/nar/gkx1253DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5814968PMC
February 2018

Brain urea increase is an early Huntington's disease pathogenic event observed in a prodromal transgenic sheep model and HD cases.

Proc Natl Acad Sci U S A 2017 12 11;114(52):E11293-E11302. Epub 2017 Dec 11.

Centre for Brain Research, School of Biological Sciences, The University of Auckland, Auckland 1010, New Zealand;

The neurodegenerative disorder Huntington's disease (HD) is typically characterized by extensive loss of striatal neurons and the midlife onset of debilitating and progressive chorea, dementia, and psychological disturbance. HD is caused by a CAG repeat expansion in the () gene, translating to an elongated glutamine tract in the huntingtin protein. The pathogenic mechanism resulting in cell dysfunction and death beyond the causative mutation is not well defined. To further delineate the early molecular events in HD, we performed RNA-sequencing (RNA-seq) on striatal tissue from a cohort of 5-y-old -line sheep expressing a human CAG-expansion cDNA transgene. Our HD sheep are a prodromal model and exhibit minimal pathology and no detectable neuronal loss. We identified significantly increased levels of the urea transporter in the striatum, along with other important osmotic regulators. Further investigation revealed elevated levels of the metabolite urea in the striatum and cerebellum, consistent with our recently published observation of increased urea in postmortem human brain from HD cases. Extending that finding, we demonstrate that postmortem human brain urea levels are elevated in a larger cohort of HD cases, including those with low-level neuropathology (Vonsattel grade 0/1). This elevation indicates increased protein catabolism, possibly as an alternate energy source given the generalized metabolic defect in HD. Increased urea and ammonia levels due to dysregulation of the urea cycle are known to cause neurologic impairment. Taken together, our findings indicate that aberrant urea metabolism could be the primary biochemical disruption initiating neuropathogenesis in HD.
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http://dx.doi.org/10.1073/pnas.1711243115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5748180PMC
December 2017

Estimation of linkage disequilibrium and effective population size in New Zealand sheep using three different methods to create genetic maps.

BMC Genet 2017 07 21;18(1):68. Epub 2017 Jul 21.

Department of Mathematics and Statistics, University of Otago, Dunedin, 9058, New Zealand.

Background: Investments in genetic selection have played a major role in the New Zealand sheep industry competitiveness. Selection may erode genetic diversity, which is a crucial factor for the success of breeding programs. Better understanding of linkage disequilibrium (LD) and ancestral effective population size (N) through quantifying this diversity and comparison between populations allows for more informed decisions with regards to selective breeding taking population genetic diversity into account. The estimation of N can be determined via genetic markers and requires knowledge of genetic distances between these markers. Single nucleotide polymorphisms (SNP) data from a sample of 12,597 New Zealand crossbred and purebred sheep genotyped with the Illumina Ovine SNP50 BeadChip was used to perform a genome-wide scan of LD and N . Three methods to estimate genetic distances were investigated: 1) M1: a ratio fixed across the whole genome of one Megabase per centiMorgan; 2) M2: the ratios of genetic distance (using M3, below) over physical distance fixed for each chromosome; and, 3) M3: a genetic map of inter-SNP distances estimated using CRIMAP software (v2.503).

Results: The estimates obtained with M2 and M3 showed much less variability between autosomes than those with M1, which tended to give lower N results and higher LD decay. The results suggest that N has decreased since the development of sheep breeds in Europe and this reduction in N has been accelerated in the last three decades. The N estimated for five generations in the past ranged from 71 to 237 for Texel and Romney breeds, respectively. A low level of genetic kinship and inbreeding was estimated in those breeds suggesting avoidance of mating close relatives.

Conclusions: M3 was considered the most accurate method to create genetic maps for the estimation of LD and N. The findings of this study highlight the history of genetic selection in New Zealand crossbred and purebred sheep and these results will be very useful to understand genetic diversity of the population with respect to genetic selection. In addition, it will help geneticists to identify genomic regions which have been preferentially selected within a variety of breeds and populations.
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http://dx.doi.org/10.1186/s12863-017-0534-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5521107PMC
July 2017

Copy number variants in the sheep genome detected using multiple approaches.

BMC Genomics 2016 06 8;17:441. Epub 2016 Jun 8.

AgResearch, Invermay Agricultural Centre, PB 50034, Mosgiel, 9053, New Zealand.

Background: Copy number variants (CNVs) are a type of polymorphism found to underlie phenotypic variation, both in humans and livestock. Most surveys of CNV in livestock have been conducted in the cattle genome, and often utilise only a single approach for the detection of copy number differences. Here we performed a study of CNV in sheep, using multiple methods to identify and characterise copy number changes. Comprehensive information from small pedigrees (trios) was collected using multiple platforms (array CGH, SNP chip and whole genome sequence data), with these data then analysed via multiple approaches to identify and verify CNVs.

Results: In total, 3,488 autosomal CNV regions (CNVRs) were identified in this study, which substantially builds on an initial survey of the sheep genome that identified 135 CNVRs. The average length of the identified CNVRs was 19 kb (range of 1 kb to 3.6 Mb), with shorter CNVRs being more frequent than longer CNVRs. The total length of all CNVRs was 67.6Mbps, which equates to 2.7 % of the sheep autosomes. For individuals this value ranged from 0.24 to 0.55 %, and the majority of CNVRs were identified in single animals. Rather than being uniformly distributed throughout the genome, CNVRs tended to be clustered. Application of three independent approaches for CNVR detection facilitated a comparison of validation rates. CNVs identified on the Roche-NimbleGen 2.1M CGH array generally had low validation rates with lower density arrays, while whole genome sequence data had the highest validation rate (>60 %).

Conclusions: This study represents the first comprehensive survey of the distribution, prevalence and characteristics of CNVR in sheep. Multiple approaches were used to detect CNV regions and it appears that the best method for verifying CNVR on a large scale involves using a combination of detection methodologies. The characteristics of the 3,488 autosomal CNV regions identified in this study are comparable to other CNV regions reported in the literature and provide a valuable and sizeable addition to the small subset of published sheep CNVs.
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http://dx.doi.org/10.1186/s12864-016-2754-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4898393PMC
June 2016

Construction of relatedness matrices using genotyping-by-sequencing data.

BMC Genomics 2015 Dec 9;16:1047. Epub 2015 Dec 9.

AgResearch, Invermay Agricultural Centre, Private Bag 50034, Mosgiel, 9053, New Zealand.

Background: Genotyping-by-sequencing (GBS) is becoming an attractive alternative to array-based methods for genotyping individuals for a large number of single nucleotide polymorphisms (SNPs). Costs can be lowered by reducing the mean sequencing depth, but this results in genotype calls of lower quality. A common analysis strategy is to filter SNPs to just those with sufficient depth, thereby greatly reducing the number of SNPs available. We investigate methods for estimating relatedness using GBS data, including results of low depth, using theoretical calculation, simulation and application to a real data set.

Results: We show that unbiased estimates of relatedness can be obtained by using only those SNPs with genotype calls in both individuals. The expected value of this estimator is independent of the SNP depth in each individual, under a model of genotype calling that includes the special case of the two alleles being read at random. In contrast, the estimator of self-relatedness does depend on the SNP depth, and we provide a modification to provide unbiased estimates of self-relatedness. We refer to these methods of estimation as kinship using GBS with depth adjustment (KGD). The estimators can be calculated using matrix methods, which allow efficient computation. Simulation results were consistent with the methods being unbiased, and suggest that the optimal sequencing depth is around 2-4 for relatedness between individuals and 5-10 for self-relatedness. Application to a real data set revealed that some SNP filtering may still be necessary, for the exclusion of SNPs which did not behave in a Mendelian fashion. A simple graphical method (a 'fin plot') is given to illustrate this issue and to guide filtering parameters.

Conclusion: We provide a method which gives unbiased estimates of relatedness, based on SNPs assayed by GBS, which accounts for the depth (including zero depth) of the genotype calls. This allows GBS to be applied at read depths which can be chosen to optimise the information obtained. SNPs with excess heterozygosity, often due to (partial) polyploidy or other duplications can be filtered based on a simple graphical method.
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http://dx.doi.org/10.1186/s12864-015-2252-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4675043PMC
December 2015

SNPchiMp v.3: integrating and standardizing single nucleotide polymorphism data for livestock species.

BMC Genomics 2015 Apr 10;16:283. Epub 2015 Apr 10.

Bioinformatics and Biostatistical Genomics group, Fondazione Parco Tecnologico Padano, Via Einstein, Loc. Cascina Codazza, 26900, Lodi, Italy.

Background: In recent years, the use of genomic information in livestock species for genetic improvement, association studies and many other fields has become routine. In order to accommodate different market requirements in terms of genotyping cost, manufacturers of single nucleotide polymorphism (SNP) arrays, private companies and international consortia have developed a large number of arrays with different content and different SNP density. The number of currently available SNP arrays differs among species: ranging from one for goats to more than ten for cattle, and the number of arrays available is increasing rapidly. However, there is limited or no effort to standardize and integrate array- specific (e.g. SNP IDs, allele coding) and species-specific (i.e. past and current assemblies) SNP information.

Results: Here we present SNPchiMp v.3, a solution to these issues for the six major livestock species (cow, pig, horse, sheep, goat and chicken). Original data was collected directly from SNP array producers and specific international genome consortia, and stored in a MySQL database. The database was then linked to an open-access web tool and to public databases. SNPchiMp v.3 ensures fast access to the database (retrieving within/across SNP array data) and the possibility of annotating SNP array data in a user-friendly fashion.

Conclusions: This platform allows easy integration and standardization, and it is aimed at both industry and research. It also enables users to easily link the information available from the array producer with data in public databases, without the need of additional bioinformatics tools or pipelines. In recognition of the open-access use of Ensembl resources, SNPchiMp v.3 was officially credited as an Ensembl E!mpowered tool. Availability at http://bioinformatics.tecnoparco.org/SNPchimp.
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http://dx.doi.org/10.1186/s12864-015-1497-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4399246PMC
April 2015

Mutations in the leptin receptor gene associated with delayed onset of puberty are also associated with decreased ovulation and lambing rates in prolific Davisdale sheep.

Reprod Fertil Dev 2015 Feb 18. Epub 2015 Feb 18.

The aim of this study was to determine if single nucleotide polymorphisms (SNPs) in the leptin receptor (LEPR) gene associated with delayed onset of puberty are associated with changes in other reproductive traits in adult ewes. The ovulation rate of ewes homozygous for the SNPs was ~15% lower (PPLEPR SNPs than their wild-type or heterozygous contemporaries. Partial failure of multiple ovulations was also increased (PLEPR had on average 0.2 fewer lambs at mid-pregnancy and at birth compared with the wild-type or heterozygous ewes (PLEPR were strongly associated with poorer reproductive performance in Davisdale ewes, which is likely to be linked to both a reduced number of ova available for fertilisation and an increased number of ewes failing to become pregnant. Increased partial failure of multiple ovulations in ewes with high ovulation rates (i.e. 3 or greater) may also contribute to the poor reproductive performance.
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http://dx.doi.org/10.1071/RD14382DOI Listing
February 2015

The accuracy, feasibility and challenges of sequencing short tandem repeats using next-generation sequencing platforms.

PLoS One 2014 1;9(12):e113862. Epub 2014 Dec 1.

Department of Anatomy, University of Otago, Dunedin, New Zealand; Allan Wilson Centre for Molecular Ecology and Evolution, University of Otago, Dunedin, New Zealand.

To date we have little knowledge of how accurate next-generation sequencing (NGS) technologies are in sequencing repetitive sequences beyond known limitations to accurately sequence homopolymers. Only a handful of previous reports have evaluated the potential of NGS for sequencing short tandem repeats (microsatellites) and no empirical study has compared and evaluated the performance of more than one NGS platform with the same dataset. Here we examined yeast microsatellite variants from both long-read (454-sequencing) and short-read (Illumina) NGS platforms and compared these to data derived through Sanger sequencing. In addition, we investigated any locus-specific biases and differences that might have resulted from variability in microsatellite repeat number, repeat motif or type of mutation. Out of 112 insertion/deletion variants identified among 45 microsatellite amplicons in our study, we found 87.5% agreement between the 454-platform and Sanger sequencing in frequency of variant detection after Benjamini-Hochberg correction for multiple tests. For a subset of 21 microsatellite amplicons derived from Illumina sequencing, the results of short-read platform were highly consistent with the other two platforms, with 100% agreement with 454-sequencing and 93.6% agreement with the Sanger method after Benjamini-Hochberg correction. We found that the microsatellite attributes copy number, repeat motif and type of mutation did not have a significant effect on differences seen between the sequencing platforms. We show that both long-read and short-read NGS platforms can be used to sequence short tandem repeats accurately, which makes it feasible to consider the use of these platforms in high-throughput genotyping. It appears the major requirement for achieving both high accuracy and rare variant detection in microsatellite genotyping is sufficient read depth coverage. This might be a challenge because each platform generates a consistent pattern of non-uniform sequence coverage, which, as our study suggests, may affect some types of tandem repeats more than others.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0113862PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4250034PMC
September 2015

Linkage disequilibrium over short physical distances measured in sheep using a high-density SNP chip.

Anim Genet 2014 Oct 17;45(5):754-7. Epub 2014 Jul 17.

CSIRO Agriculture Flagship, St Lucia, Qld 4067, Australia.

The extent of linkage disequilibrium (LD) between genetic loci has implications for both association studies and the accuracy of genomic prediction. To characterise the persistence of LD in diverse sheep breeds, two SNP genotyping platforms were used. First, existing SNP genotypes from 63 breeds obtained using the ovine SNP50 BeadChip (49,034 loci) were used to estimate LD decay in populations with contrasting levels of genetic diversity. Given the paucity of marker pairs separated by short physical distances on the SNP50 BeadChip, genotyping was subsequently performed for four breeds using the recently developed ovine HD BeadChip that assays approximately 600,000 SNPs with an average genomic spacing of 5 kb. This facilitated a highly accurate estimate of LD over short genomic distances (<30 kb) and revealed LD varies considerably between sheep breeds. Further, sheep appear to contain generally lower levels of LD than do other domestic species, likely a reflection of aspects of their past population history.
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http://dx.doi.org/10.1111/age.12197DOI Listing
October 2014

Genome-wide DNA methylation patterns and transcription analysis in sheep muscle.

PLoS One 2014 10;9(7):e101853. Epub 2014 Jul 10.

AgResearch Ltd., Invermay Agricultural Centre, Puddle Alley, Mosgiel, New Zealand.

DNA methylation plays a central role in regulating many aspects of growth and development in mammals through regulating gene expression. The development of next generation sequencing technologies have paved the way for genome-wide, high resolution analysis of DNA methylation landscapes using methodology known as reduced representation bisulfite sequencing (RRBS). While RRBS has proven to be effective in understanding DNA methylation landscapes in humans, mice, and rats, to date, few studies have utilised this powerful method for investigating DNA methylation in agricultural animals. Here we describe the utilisation of RRBS to investigate DNA methylation in sheep Longissimus dorsi muscles. RRBS analysis of ∼1% of the genome from Longissimus dorsi muscles provided data of suitably high precision and accuracy for DNA methylation analysis, at all levels of resolution from genome-wide to individual nucleotides. Combining RRBS data with mRNAseq data allowed the sheep Longissimus dorsi muscle methylome to be compared with methylomes from other species. While some species differences were identified, many similarities were observed between DNA methylation patterns in sheep and other more commonly studied species. The RRBS data presented here highlights the complexity of epigenetic regulation of genes. However, the similarities observed across species are promising, in that knowledge gained from epigenetic studies in human and mice may be applied, with caution, to agricultural species. The ability to accurately measure DNA methylation in agricultural animals will contribute an additional layer of information to the genetic analyses currently being used to maximise production gains in these species.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0101853PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4092064PMC
March 2015

The sheep genome illuminates biology of the rumen and lipid metabolism.

Science 2014 Jun;344(6188):1168-1173

Human Genome Sequencing Center, Baylor College of Medicine, Houston, TX 77030, USA.

Sheep (Ovis aries) are a major source of meat, milk, and fiber in the form of wool and represent a distinct class of animals that have a specialized digestive organ, the rumen, that carries out the initial digestion of plant material. We have developed and analyzed a high-quality reference sheep genome and transcriptomes from 40 different tissues. We identified highly expressed genes encoding keratin cross-linking proteins associated with rumen evolution. We also identified genes involved in lipid metabolism that had been amplified and/or had altered tissue expression patterns. This may be in response to changes in the barrier lipids of the skin, an interaction between lipid metabolism and wool synthesis, and an increased role of volatile fatty acids in ruminants compared with nonruminant animals.
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http://dx.doi.org/10.1126/science.1252806DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4157056PMC
June 2014

Single-nucleotide polymorphisms in the LEPR gene are associated with divergent phenotypes for age at onset of puberty in Davisdale ewes.

Biol Reprod 2014 Feb 20;90(2):33. Epub 2014 Feb 20.

Animal Reproduction Division, Indian Council of Agriculture Research Complex for North Eastern Hill Region, Tripura Centre, Lembucherra, West Tripura, India.

Attainment of puberty is a key developmental event influenced by genetic and environmental factors. In examining age at attainment of puberty, we observed closely related rams from the Davisdale line whose daughters differed in age at which they attained puberty. A candidate gene approach was used to identify mutations that may underlie these observed differences. Four rams with divergent phenotypes for their daughter's age at onset of puberty were selected for whole-genome sequencing. The coding regions of genes with known roles in regulating reproductive function were searched for single-nucleotide polymorphisms (SNPs) that altered the amino acid sequence of the protein. Of interest were three SNPs in the leptin receptor gene (LEPR). A Sequenom assay was developed to determine the genotype of these SNPs in daughters of 17 sons of a founding sire. A higher percentage of ewe lambs homozygous for the LEPR mutations failed to undergo puberty before 1 yr of age, and those that did undergo puberty during the first breeding season on average were approximately 17 days older than homozygous wild-type ewes. Heterozygous ewes were intermediate for both measurements. Given the predicted change in protein function produced by the mutation in LEPR and the strong associations between the genotype and onset of puberty phenotypes, we propose that this mutation in LEPR underlies the observed difference in age at onset of puberty in the Davisdale line. Furthermore, these animals will likely provide a useful model to better understand the role of leptin in the regulation of puberty.
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http://dx.doi.org/10.1095/biolreprod.113.115923DOI Listing
February 2014

Development of a predicted physical map of microsatellite locus positions for pinnipeds, with wider applicability to the Carnivora.

Mol Ecol Resour 2011 May 28;11(3):503-13. Epub 2010 Dec 28.

Department of Anatomy and Structural Biology, Centre for Reproduction and Genomics, University of Otago, Dunedin, New Zealand.

Understanding genetic variation responsible for phenotypic differences in natural populations is significantly hampered by a lack of genomic data for many species. Levels of variation can, however, be estimated using microsatellite markers, which may be useful for relating individual fitness to genetic diversity. Prior studies have demonstrated correlations between heterozygosity and individual fitness in some species. These correlations are sometimes driven by a subset of markers, and it is unclear whether this is because those markers best reflect genome-wide heterozygosity, or whether they are linked to fitness-related genes. Differentiating between these scenarios is hindered when the genomic location of markers is unknown. Here, we develop a predicted genomic map of pinniped microsatellite loci based on conservation of primary sequence and genomic location between dog, cat and giant panda. We mapped 210 of 260 (81%) microsatellites from pinnipeds to locations in dog, cat and giant panda genomes. Based on the demonstrable synteny between the genomes of closely related taxa within the Carnivora, we use these data to identify those microsatellites with the greatest chance of cross-species amplification success and demonstrate successful amplification of 21 of 26 loci for cat, dog and two seal species. We also demonstrate the potential to identify candidate genes that may underpin the functional relationship with individual fitness. Overall, we show that this approach provides a rapid and robust method to elucidate genome organisation for nonmodel organisms and have established a resource that facilitates further genetic research on pinnipeds that also has wider applicability to other carnivores.
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http://dx.doi.org/10.1111/j.1755-0998.2010.02962.xDOI Listing
May 2011

Resolving the evolution of extant and extinct ruminants with high-throughput phylogenomics.

Proc Natl Acad Sci U S A 2009 Nov 21;106(44):18644-9. Epub 2009 Oct 21.

Divisions of Animal Sciences, Biological Sciences, and Department of Veterinary Pathobiology, University of Missouri, Columbia, MO 65211, USA.

The Pecorans (higher ruminants) are believed to have rapidly speciated in the Mid-Eocene, resulting in five distinct extant families: Antilocapridae, Giraffidae, Moschidae, Cervidae, and Bovidae. Due to the rapid radiation, the Pecoran phylogeny has proven difficult to resolve, and 11 of the 15 possible rooted phylogenies describing ancestral relationships among the Antilocapridae, Giraffidae, Cervidae, and Bovidae have each been argued as representations of the true phylogeny. Here we demonstrate that a genome-wide single nucleotide polymorphism (SNP) genotyping platform designed for one species can be used to genotype ancient DNA from an extinct species and DNA from species diverged up to 29 million years ago and that the produced genotypes can be used to resolve the phylogeny for this rapidly radiated infraorder. We used a high-throughput assay with 54,693 SNP loci developed for Bos taurus taurus to rapidly genotype 678 individuals representing 61 Pecoran species. We produced a highly resolved phylogeny for this diverse group based upon 40,843 genome-wide SNP, which is five times as many informative characters as have previously been analyzed. We also establish a method to amplify and screen genomic information from extinct species, and place Bison priscus within the Bovidae. The quality of genotype calls and the placement of samples within a well-supported phylogeny may provide an important test for validating the fidelity and integrity of ancient samples. Finally, we constructed a phylogenomic network to accurately describe the relationships between 48 cattle breeds and facilitate inferences concerning the history of domestication and breed formation.
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http://dx.doi.org/10.1073/pnas.0904691106DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2765454PMC
November 2009

A multiway analysis for identifying high integrity bovine BACs.

BMC Genomics 2009 Jan 23;10:46. Epub 2009 Jan 23.

CSIRO Livestock Industries, 306 Carmody Road, St. Lucia, QLD 4067, Australia.

Background: In large genomics projects involving many different types of analyses of bacterial artificial chromosomes (BACs), such as fingerprinting, end sequencing (BES) and full BAC sequencing there are many opportunities for the identities of BACs to become confused. However, by comparing the results from the different analyses, inconsistencies can be identified and a set of high integrity BACs preferred for future research can be defined.

Results: The location of each bovine BAC in the BAC fingerprint-based genome map and in the genome assembly were compared based on the reported BESs, and for a smaller number of BACs the full sequence. BACs with consistent positions in all three datasets, or if the full sequence was not available, for both the fingerprint map and BES-based alignments, were deemed to be correctly positioned. BACs with consistent BES-based and fingerprint-based locations, but with conflicting locations based on the fully sequenced BAC, appeared to have been misidentified during sequencing, and included a number of apparently swapped BACs. Inconsistencies between BES-based and fingerprint map positions identified thirty one plates from the CHORI-240 library that appear to have suffered substantial systematic problems during the end-sequencing of the BACs. No systematic problems were identified in the fingerprinting of the BACs. Analysis of BACs overlapping in the assembly identified a small overrepresentation of clones with substantial overlap in the library and a substantial enrichment of highly overlapping BACs on the same plate in the CHORI-240 library. More than half of these BACs appear to have been present as duplicates on the original BAC-library plates and thus should be avoided in subsequent projects.

Conclusion: Our analysis shows that approximately 95% of the bovine CHORI-240 library clones with both a BAC fingerprint and two BESs mapping to the genome in the expected orientations (approximately 27% of all BACs) have consistent locations in the BAC fingerprint map and the genome assembly. We have developed a broadly applicable methodology for checking the integrity of BAC-based datasets even where only incomplete and partially assembled genomic sequence is available.
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http://dx.doi.org/10.1186/1471-2164-10-46DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2660975PMC
January 2009

A physical map of the bovine genome.

Genome Biol 2007 ;8(8):R165

USDA, ARS, US Meat Animal Research Center, Clay Center, NE 68933, USA.

Background: Cattle are important agriculturally and relevant as a model organism. Previously described genetic and radiation hybrid (RH) maps of the bovine genome have been used to identify genomic regions and genes affecting specific traits. Application of these maps to identify influential genetic polymorphisms will be enhanced by integration with each other and with bacterial artificial chromosome (BAC) libraries. The BAC libraries and clone maps are essential for the hybrid clone-by-clone/whole-genome shotgun sequencing approach taken by the bovine genome sequencing project.

Results: A bovine BAC map was constructed with HindIII restriction digest fragments of 290,797 BAC clones from animals of three different breeds. Comparative mapping of 422,522 BAC end sequences assisted with BAC map ordering and assembly. Genotypes and pedigree from two genetic maps and marker scores from three whole-genome RH panels were consolidated on a 17,254-marker composite map. Sequence similarity allowed integrating the BAC and composite maps with the bovine draft assembly (Btau3.1), establishing a comprehensive resource describing the bovine genome. Agreement between the marker and BAC maps and the draft assembly is high, although discrepancies exist. The composite and BAC maps are more similar than either is to the draft assembly.

Conclusion: Further refinement of the maps and greater integration into the genome assembly process may contribute to a high quality assembly. The maps provide resources to associate phenotypic variation with underlying genomic variation, and are crucial resources for understanding the biology underpinning this important ruminant species so closely associated with humans.
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http://dx.doi.org/10.1186/gb-2007-8-8-r165DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2374996PMC
February 2008

PATH: a task for the inference of phylogenies.

Bioinformatics 2002 Apr;18(4):646-7

Department of Molecular Biophysics, DKFZ, INF280, D-69120 Heidelberg, Germany.

Unlabelled: Phylogenetic Analysis Task in Husar (PATH) is a task for the inference of phylogenies. It executes three phylogenetic methods and automatically chooses the evolutionary model for each set of data. The output of the tasks shows the consensus trees together with full results obtained from all executed methods.

Availability: PATH is available at the German EMBnet node after registration via www at http://genome.dkfz-heidelberg.de
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http://dx.doi.org/10.1093/bioinformatics/18.4.646DOI Listing
April 2002
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