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Abstract Analysis of the genetic variation of an endangered population is an important component for the success of conservation. Animals from two local Romanian pig breeds, the Mangalitsa and Bazna, were analysed for variation at a number of genetic loci using PCR-based DNA tests. Polymorphism was assessed at loci which 1) are known to cause phenotypic variation, 2) are potentially involved in trait differences or 3) are putative candidate genes. The traits considered are disease resistance, growth, coat colour, meat quality and prolificacy. Even though the populations are small and the markers are limited to specific genes, we found significant differences in five of the ten characterised loci. In some cases the observed allele frequencies were interesting in relation to gene function and the phenotype of the breed. These breeds are part of a conservation programme in Romania and marker information may be useful in preserving a representative gene pool in the populations. The use of polymorphisms in type 1 (gene) markers may be a useful complement to analysis based on anonymous markers.

Abstract The accurate estimation of the probability of identity by descent (IBD) at loci or genome positions of interest is paramount to the genetic study of quantitative and disease resistance traits. We present a Monte Carlo Markov Chain method to compute IBD probabilities between individuals conditional on DNA markers and on pedigree information. The IBDs can be obtained in a completely general pedigree at any genome position of interest, and all marker and pedigree information available is used. The method can be split into two steps at each iteration. First, phases are sampled using current genotypic configurations of relatives and second, crossover events are simulated conditional on phases. Internal track is kept of all founder origins and crossovers such that the IBD probabilities averaged over replicates are rapidly obtained. We illustrate the method with some examples. First, we show that all pedigree information should be used to obtain line origin probabilities in F2 crosses. Second, the distribution of genetic relationships between half and full sibs is analysed in both simulated data and in real data from an F2 cross in pigs.

Abstract Background Understanding the role of host genetics in resistance to porcine reproductive and respiratory syndrome virus (PRRSV) infection, and the effects of PRRS on pig health and related growth, are goals of the PRRS Host Genetics Consortium (PHGC). Methods The project uses a nursery pig model to assess pig resistance/susceptibility to primary PRRSV infection. To date, 6 groups of 200 crossbred pigs from high health farms were donated by commercial sources. After acclimation, the pigs were infected with PRRSV in a biosecure facility and followed for 42 days post infection (dpi). Blood samples were collected at 0, 4, 7, 10, 14, 21, 28, 35 and 42 dpi for serum and whole blood RNA gene expression analyses; weekly weights were recorded for growth traits. All data have been entered into the PHGC relational database. Genomic DNAs from all PHGC1-6 pigs were prepared and genotyped with the Porcine SNP60 SNPchip. Results Results have affirmed that all challenged pigs become PRRSV infected with peak viremia being observed between 4-21 dpi. Multivariate statistical analyses of viral load and weight data have identified PHGC pigs in different virus/weight categories. Sera are now being compared for factors involved in recovery from infection, including speed of response and levels of immune cytokines. Genome-wide association studies (GWAS) are underway to identify genes and chromosomal locations that identify PRRS resistant/susceptible pigs and pigs able to maintain growth while infected with PRRSV. Conclusions Overall, the PHGC project will enable researchers to discover and verify important genotypes and phenotypes that predict resistance/susceptibility to PRRSV infection. The availability of PHGC samples provides a unique opportunity to continue to develop deeper phenotypes on every PRRSV infected pig.

Tabby patterns of fur coats are defining characteristics in wild and domestic felids. Historically, three autosomal alleles at one locus (Tabby): Abyssinian (Ta ; a.k.a. ticked), mackerel (Tm ; a.k.a. striped) and blotched (tb ; a.k.a. classic, blotched) were thought to control these patterns in domestic cats and their breeds. Currently, at least three loci influence cat tabby markings, two of which are designated Tabby and Ticked. The Tabby locus is laeverin (LVRN) and affects the mackerel and blotched patterns. The unidentified gene for the Ticked locus on cat chromosome B1 was suggested to control the presence or absence of the ticked pattern (Tabby - Abyssinian (Ta ; a.k.a. ticked). The cat reference genome (Cinnamon, the Abyssinian) has the ticked phenotype and the variant dataset and coat phenotypes from the 99 Lives Cat Genome Consortium (195 cats) were used to identify candidate genes and variants associated with the Ticked locus. Two strategies were used to find the Ticked allele(s), one considered Cinnamon with the reference allele or heterozygous (Strategy A) and the other considered Cinnamon as having the variant allele or heterozygous (Strategy B). For Strategy A, two variants in Dickkopf Wnt Signaling Pathway Inhibitor 4 (DKK4), a p.Cys63Tyr (B1:41621481, c.188G>A) and a less common p.Ala18Val (B1:42620835, c.53C>T) variant are suggested as two alleles influencing the Ticked phenotype. Bioinformatic and molecular modeling analysis suggests that these changes disrupt a key disulfide bond in the Dkk4 cysteine-rich domain 1 or Dkk4 signal peptide cleavage respectively. All coding variants were excluded as Ticked alleles using Strategy B.

Abstract Background The pig genome is being sequenced and characterised under the auspices of the Swine Genome Sequencing Consortium. The sequencing strategy followed a hybrid approach combining hierarchical shotgun sequencing of BAC clones and whole genome shotgun sequencing. Results Assemblies of the BAC clone derived genome sequence have been annotated using the Pre-Ensembl and Ensembl automated pipelines and made accessible through the Pre-Ensembl/Ensembl browsers. The current annotated genome assembly (Sscrofa9) was released with Ensembl 56 in September 2009. A revised assembly (Sscrofa10) is under construction and will incorporate whole genome shotgun sequence (WGS) data providing > 30× genome coverage. The WGS sequence, most of which comprise short Illumina/Solexa reads, were generated from DNA from the same single Duroc sow as the source of the BAC library from which clones were preferentially selected for sequencing. In accordance with the Bermuda and Fort Lauderdale agreements and the more recent Toronto Statement the data have been released into public sequence repositories (Genbank/EMBL, NCBI/Ensembl trace repositories) in a timely manner and in advance of publication. Conclusions In this marker paper, the Swine Genome Sequencing Consortium (SGSC) sets outs its plans for analysis of the pig genome sequence, for the application and publication of the results.

Abstract Background Identification and culling of replacement gilts with poor skeletal conformation and feet and leg (FL) unsoundness is an approach used to reduce sow culling and mortality rates in breeding stock. Few candidate genes related to soundness traits have been identified in the pig. Methods In this study, 2066 commercial females were scored for 17 traits describing body conformation and FL structure, and were used for association analyses. Genotyping of 121 SNPs derived from 95 genes was implemented using Sequenom's MassARRAY system. Results Based on the association results from single trait and principal components using mixed linear model analyses and false discovery rate testing, it was observed that APOE, BMP8, CALCR, COL1A2, COL9A1, DKFZ, FBN1 and VDBP were very highly significantly (P < 0.001) associated with body conformation traits. The genes ALOX5, BMP8, CALCR, OPG, OXTR and WNT16 were very highly significantly (P < 0.001) associated with FL structures, and APOE, CALCR, COL1A2, GNRHR, IHH, MTHFR and WNT16 were highly significantly (P < 0.01) associated with overall leg action. Strong linkage disequilibrium between CALCR and COL1A2 on SSC9 was detected, and haplotype -ACGACC- was highly significantly (P < 0.01) associated with overall leg action and several important FL soundness traits. Conclusion The present findings provide a comprehensive list of candidate genes for further use in fine mapping and biological functional analyses.

Abstract Background MicroRNAs (miRNAs) are recognized as one of the most important families of non-coding RNAs that serve as important sequence-specific post-transcriptional regulators of gene expression. Identification of miRNAs is an important requirement for understanding the mechanisms of post-transcriptional regulation. Hundreds of miRNAs have been identified by direct cloning and computational approaches in several species. However, there are still many miRNAs that remain to be identified due to lack of either sequence features or robust algorithms to efficiently identify them. Results We have evaluated features valuable for pre-miRNA prediction, such as the local secondary structure differences of the stem region of miRNA and non-miRNA hairpins. We have also established correlations between different types of mutations and the secondary structures of pre-miRNAs. Utilizing these features and combining some improvements of the current pre-miRNA prediction methods, we implemented a computational learning method SVM (support vector machine) to build a high throughput and good performance computational pre-miRNA prediction tool called MiRFinder. The tool was designed for genome-wise, pair-wise sequences from two related species. The method built into the tool consisted of two major steps: 1) genome wide search for hairpin candidates and 2) exclusion of the non-robust structures based on analysis of 18 parameters by the SVM method. Results from applying the tool for chicken/human and D. melanogaster/D. pseudoobscura pair-wise genome alignments showed that the tool can be used for genome wide pre-miRNA predictions. Conclusion The MiRFinder can be a good alternative to current miRNA discovery software. This tool is available at http://www.bioinformatics.org/mirfinder/.

State-of-the-art health care includes genome sequencing of the patient to identify genetic variants that contribute to either the cause of their malady or variants that can be targeted to improve treatment. The goal was to introduce state-of-the-art health care to cats using genomics and a precision medicine approach. To test the feasibility of a precision medicine approach in domestic cats, a single cat that presented to the University of Missouri, Veterinary Health Center with an undiagnosed neurologic disease was whole-genome sequenced. The DNA variants from the cat were compared to the DNA variant database produced by the 99 Lives Cat Genome Sequencing Consortium. Approximately 25× genomic coverage was produced for the cat. A predicted p.H441P missense mutation was identified in NPC1, the gene causing Niemann-Pick type C1 on cat chromosome D3.47456793 caused by an adenine-to-cytosine transversion, c.1322A>C. The cat was homozygous for the variant. The variant was not identified in any other 73 domestic and 9 wild felids in the sequence database or 190 additionally genotyped cats of various breeds. The successful effort suggested precision medicine is feasible for cats and other undiagnosed cats may benefit from a genomic analysis approach. The 99 Lives DNA variant database was sufficient but would benefit from additional cat sequences. Other cats with the mutation may be identified and could be introduced as a new biomedical model for NPC1. A genetic test could eliminate the disease variant from the population.