Excess Homozygosity Research Articles

Assessment of Genetic Diversity in Azerbaijani Buffalo Population in Iran Based on Runs of Homozygosity Stretches

Quantifying human, plant and livestock inbreeding has been an important goal of evolutionary biologists and agricultural scientists for nearly a century. Comprehensive identification of the breed characteristics is a critical factor for the effective management of farm animal genetic resources. The objective of this study was to estimate inbreeding coefficients derived from runs of homozygosity islands from 243 Iranian Azerbaijani buffalos using dense SNP markers (Axiom® Buffalo Genotyping 90K). After quality control, 62,141 SNP markers remained for computation of inbreeding coefficients. The amount of estimated inbreeding coefficient using four methods including genomic relationship matrix (FGRM), excess of homozygosity (FHOM), correlation of uniting gametes (FUNI) and FIS was similar (0.026) and it was estimated 0.0163 using Run of homozygosity procedure. Low levels of FROH in Azerbaijan buffalo population can be due to the traditional rural breeding system, random mating, high recombination rate and small population of ancestors.

Fine-scale estimation of inbreeding rates, runs of homozygosity and genome-wide heterozygosity levels in the Mangalarga Marchador horse breed.

With the availability of high-density SNP panels and the establishment of approaches for characterizing homozygosity and heterozygosity sites, it is possible to access fine-scale information regarding genomes, providing more than just comparisons of different inbreeding coefficients. This is the first study that seeks to access such information for the Mangalarga Marchador (MM) horse breed on a genomic scale. To this end, we aimed to assess inbreeding levels using different coefficients, as well as to characterize homozygous and heterozygous runs in the population. Using Axiom ® Equine Genotyping Array-670k SNP (Thermo Fisher), 192 horses were genotyped. Our results showed different estimates: inbreeding from genomic coefficients (FROH )=0.16; pedigree-based (FPED )=0.008; and a method based on excess homozygosity (FHOM )=0.010. The correlations between the inbreeding coefficients were low to moderate, and some comparisons showed negative correlations, being practically null. In total, 85,295 runs of homozygosity (ROH) and 10,016 runs of heterozygosity (ROHet) were characterized for the 31 horse autosomal chromosomes. The class with the highest percentage of ROH was 0-2Mbps, with 92.78% of the observations. In the ROHet results, only the 0-2 class presented observations, with chromosome 11 highlighted in a region with high genetic variability. Three regions from the ROHet analyses showed genes with known functions: tripartite motif-containing 37 (TRIM37), protein phosphatase, Mg2+ /Mn2+ dependent 1E (PPM1E) and carbonic anhydrase 10 (CA10). Therefore, our findings suggest moderate inbreeding, possibly attributed to breed formation, annulling possible recent inbreeding. Furthermore, regions with high variability in the MM genome were identified (ROHet), associated with the recent selection and important events in the development and performance of MM horses over generations.

Detection of selection signatures in Limousin cattle using whole-genome resequencing.

Limousin, a renowned beef breed originating from central France, has been selectively bred over the last 100years to improve economically important traits. We used whole-genome sequencing data from 10 unrelated Limousin bull calves to detect polymorphisms and identify regions under selection. A total of 13943766 variants were identified. Moreover, 311852 bi-allelic SNPs and 92229 indels located on autosomes were fixed for the alternative allele in all sequenced animals, including the previously reported missense deleterious F94L mutation in MSTN. We performed a whole-genome screen to discover genomic regions with excess homozygosity, using the pooled heterozygosity score and identified 171 different candidate selective sweeps. In total, 68 candidate genes were found in only 57 of these regions, indicating that a large fraction of the genome under selection might lie in non-coding regions and suggesting that a majority of adaptive mutations might be regulatory in nature. Many QTL were found within candidate selective sweep regions, including QTL associated with shear force or carcass weight. Among the putative selective sweeps, we located genes (MSTN, NCKAP5, RUNX2) that potentially contribute to important phenotypes in Limousin. Several candidate regions and genes under selection were also found in previous genome-wide selection scans performed in Limousin. In addition, we were able to pinpoint candidate causative regulatory polymorphisms in GRIK3 and RUNX2 that might have been under selection. Our results will contribute to improved understanding of the mechanisms and targets of artificial selection and will facilitate the interpretation of GWASs performed in Limousin.

Microsatellite Genotyping Corroborated Loss of Genetic Diversity in Clarias batrachus as a Result of Lack of Regulatory Reforms in Aquaculture.

In India, over the past 50 years, aquaculture practices of species such as those used for Clarias batrachus were developed without adequate regulatory oversight. In these situations, it is important to consider the influence that genetic factors can have on such vulnerable aquaculture species. Population genetic structure can be evaluated through the use of neutral molecular markers, and this can aid in predicting the risk of the demise of populations and for framing management strategies to conserve remaining populations. The study presented here reports on the genetic status of C. batrachus populations through the analysis of data collected using 22 microsatellite markers from seven natural and one hatchery population. The mean values for observed heterozygosity across loci within populations ranged from 0.242 to 0.485. Measures of genetic differentiation were low overall, with mean values for FST of 0.270, FIS of 0.113 and FIT of 0.353. An AMOVA analysis revealed that percentages of variation among and within populations were 27.16 and 6.86, respectively, and Bayesian clustering analyses showed a population subdivision consisting of five clusters with admixture of haplotypes from other populations leading to genetic bottleneck. We also examined how hatchery management factors leading to excessive exchanges of fish between river systems through could impact the structure of the C. batrachus populations. Overall, this study shows how the systematic use of molecular markers can facilitate the development of management policies for these populations and for the development of a comprehensive set of rules for hatcheries and aquaculture practices, including avoidance of excessive homozygosity by avoiding repeated use of feral broodstock and their interrogation.

Linkage Disequilibrium, Effective Population Size and Genomic Inbreeding Rates in American Mink Using Genotyping-by-Sequencing Data.

Knowledge of linkage disequilibrium (LD) patterns is necessary to determine the minimum density of markers required for genomic studies and to infer historical changes as well as inbreeding events in the populations. In this study, we used genotyping-by-sequencing (GBS) approach to detect single nucleotide polymorphisms (SNPs) across American mink genome and further to estimate LD, effective population size (Ne), and inbreeding rates based on excess of homozygosity (FHOM) and runs of homozygosity (ROH). A GBS assay was constructed based on the sequencing of ApeKI-digested libraries from 285 American mink using Illumina HiSeq Sequencer. Data of 13,321 SNPs located on 46 scaffolds was used to perform LD analysis. The average LD (r2 ± SD) between adjacent SNPs was 0.30 ± 0.35 over all scaffolds with an average distance of 51 kb between markers. The average r2 < 0.2 was observed at inter-marker distances of >40 kb, suggesting that at least 60,000 informative SNPs would be required for genomic selection in American mink. The Ne was estimated to be 116 at five generations ago. In addition, the most rapid decline of population size was observed between 100 and 200 generations ago. Our results showed that short extensions of homozygous genotypes (500 kb to 1 Mb) were abundant across the genome and accounted for 33% of all ROH identified. The average inbreeding coefficient based on ROH longer than 1 Mb was 0.132 ± 0.042. The estimations of FHOM ranged from −0.44 to 0.34 among different samples with an average of 0.15 over all individuals. This study provided useful insights to determine the density of SNP panel providing enough statistical power and accuracy in genomic studies of American mink. Moreover, these results confirmed that GBS approach can be considered as a useful tool for genomic studies in American mink.

Runs of Homozygosity and NetView analyses provide new insight into the genome-wide diversity and admixture of three German cattle breeds.

Angler (RVA) and Red-and-White dual-purpose (RDN) cattle were in the past decades crossed with influential Red Holstein (RH) sires. However, genome-wide diversity studies in these breeds are lacking. The objective of the present study was to elucidate the genome-wide diversity and population structure of the three German cattle breeds. Using 40,851 single nucleotide polymorphism markers scored in 337 individuals, runs of homozygosity (ROH) were analysed in each breed. Clustering and a high-resolution network visualisation analyses were performed on an extended dataset that included 11 additional (outgroup) breeds. Genetic diversity levels were high with observed heterozygosity above 0.35 in all three breeds. Only RVA had a recent past effective population size (Ne) estimate above 100 at 5 generations ago. ROH length distribution followed a similar pattern across breeds and the majority of ROH were found in the length class of >5 to 10 Mb. Estimates of average inbreeding calculated from ROH (FROH) were 0.021 (RVA), 0.045 (RDN) and 0.053 (RH). Moderate to high positive correlations were found between FROH and pedigree inbreeding (FPED) and between FROH and inbreeding derived from the excess of homozygosity (FHOM), while the intercept of the regression of FROH on FPED was above zero. The population structure analysis showed strong evidence of admixture between RVA and RH. Introgression of RDN with RH genes was minimally detected and for the first time, the study uncovered Norwegian Red Cattle ancestry in RVA. Highly heterogeneous genetic background was found for RVA and RH and as expected, the breeds of the extended dataset effectively differentiated mostly based on geographical origin, validating our findings. The results of this study confirm the impact of RH sires on RVA and RDN populations. Furthermore, a close monitoring is suggested to curb further reduction of Ne in the breeds.

Genome-wide estimation of inbreeding coefficient, effective population size and haplotype blocks in Vrindavani crossbred cattle strain of India

ABSTRACT In the present study, to analyze the genomic ROH fragments, haplotypes, inbreeding coefficient and effective population size (N e ), SNP50K data of 72 Vrindavani crossbred cattle strain were obtained. After a quality check, 24,449 biallelic SNPs were retained, which were in HWE, MAF > 0.01 and genotyping rate >95%. A total of 385 ROH were found, and chromosome 25 was mostly covered with 50.86%, while chromosome one was least with 12.42%. We classified ROH into <10Mb, 10-15Mb, 15-20Mb, >20Mb categories and observed a 3% inbreeding coefficient for the population in >20Mb category. Inbreeding from an excess of homozygosity was found to be 6% in the population. Ne was also estimated through N e Estimator and SNeP and in the recent generation, it was found to be 40 for a census size of 464 cattle. The Ne/Nc ratio was found to be 0.086 and a decreasing trend of Ne was observed for over five generations. 45.95% of the autosomal genome was covered with haplotypic structure with highest total block length in chromosome one and lowest in 28. From all the observations made in this study, it can be concluded that Vrindavani is an admixed cattle strain but have an unstable population structure in the current period.

Genetic Diversity of Ascaris in China Assessed Using Simple Sequence Repeat Markers.

The giant roundworm Ascaris infects pigs and people worldwide and causes serious diseases. The taxonomic relationship between Ascaris suum and Ascaris lumbricoides is still unclear. The purpose of the present study was to investigate the genetic diversity and population genetic structure of 258 Ascaris specimens from humans and pigs from 6 sympatric regions in Ascaris-endemic regions of China using existing simple sequence repeat data. The microsatellite markers showed a high level of allelic richness and genetic diversity in the samples. Each of the populations demonstrated excess homozygosity (Ho<He, Fis>0). According to a genetic differentiation index (Fst=0.0593), there was a high-level of gene flow in the Ascaris populations. A hierarchical analysis on molecular variance revealed remarkably high levels of variation within the populations. Moreover, a population structure analysis indicated that Ascaris populations fell into 3 main genetic clusters, interpreted as A. suum, A. lumbricoides, and a hybrid of the species. We speculated that humans can be infected with A. lumbricoides, A. suum, and the hybrid, but pigs were mainly infected with A. suum. This study provided new information on the genetic diversity and population structure of Ascaris from human and pigs in China, which can be used for designing Ascaris control strategies. It can also be beneficial to understand the introgression of host affiliation.

Quality Control of Common and Rare Variants.

Thorough data quality control (QC) is a key step to the success of high-throughput genotyping approaches. Following extensive research several criteria and thresholds have been established for data QC at the sample and variant level. Sample QC is aimed at the identification and removal (when appropriate) of individuals with (1) low call rate, (2) discrepant sex or other identity-related information, (3) excess genome-wide heterozygosity and homozygosity, (4) relations to other samples, (5) ethnicity differences, (6) batch effects, and (7) contamination. Variant QC is aimed at identification and removal or refinement of variants with (1) low call rate, (2) call rate differences by phenotypic status, (3) gross deviation from Hardy-Weinberg Equilibrium (HWE), (4) bad genotype intensity plots, (5) batch effects, (6) differences in allele frequencies with published data sets, (7) very low minor allele counts(MAC), (8) low imputation quality score, (9) low variant quality score log-odds, and (10) few or low quality reads.

Ultrasound findings provide clues to investigate founder mutations expressed as runs of homozygosity in chromosomal microarray studies.

Chromosomal microarray analysis is effectively applied prenatally to detect copy number changes. Single nucleotide polymorphism (SNP) probes included in the microarray platform can detect regions of excessive homozygosity and identical-by-descent genomic stretches. The utility of the latter as part of prenatal diagnosis is not well established. Recessive founder mutations are well recognized within distinct ethnic groups. Combining these data with prenatal sonography provides accurate focused molecular diagnoses quickly. We aimed to evaluate the application of this approach in expectant families presenting to our unit. Three unrelated gravidae presenting with specific fetal sonographic findings: (1) ventriculomegaly with encephalocele; (2) severe polyhydramnion; and (3) enlarged echogenic kidneys, underwent amniocentesis for chromosomal microarray analysis, and genome-wide human SNP array was used to analyze DNA from amniocytes. The Genomic Oligoarray and SNP array evaluation tool v3.0© was used to detect recessive loci associated with the reported clinical findings. Candidate genes were further interrogated using the Israeli National Genetic Database (INGD) and specifically searching and identifying a corresponding founder mutation within the defined ethnic group. Three fetuses from 3 distinct nuclear families in which the parents shared a similar ethnicity (either Ashkenazi or Bukharan Jews) albeit no reported consanguinity were assessed. We found no copy number changes; however, by evaluating regions of homozygosity, we were able to reveal relevant candidate gene for the specific phenotype for each fetus. Using the INGD led to targeted testing of a specific homozygous fetal mutation for which parents were found to be carriers. In the fetus with ventriculomegaly with encephalocele c.1167dupA mutation in the FKTN gene, in the fetus with severe polyhydramnion c.167ins6[TTTCCC] mutation in the BSND gene, and in the fetus with enlarged echogenic kidneys, c.3761_3762delCCinsG in the PKHD1 gene were identified. A tripartite approach integrating sonographic pathology with regions of excessive homozygosity data and INGD-based founder mutation repository yields a comprehensive streamlined approach to provide accurate genetic diagnosis and counselling within the time constraints of an ongoing pregnancy.

OP22.08: Integrating ultrasound findings with chromosomal microarray stretches of homozygosity and principles in founder populations

Chromosomal microarray (CMA) analysis is effectively applied prenatally to detect copy number changes. SNP probes included on the microarray platform can detect regions of excessive homozygosity (ROH) and identical-by-descent genomic stretches. The utility of the latter in prenatal diagnosis is not well established. Recessive founder mutations have been identified within distinct ethnic groups. Combining these data with prenatal sonography provides accurate molecular diagnoses quickly. Three gravidae presenting with specific fetal sonographic findings: 1) ventriculomegaly with encephalocele; 2) severe polyhydramnion, and 3) enlarged echogenic kidneys, underwent amniocentesis for CMA, and Genome-Wide Human SNP array was used to analyse DNA from amniocytes. The Genomic Oligoarray and SNP array evaluation tool v3.0 was used to detect recessive loci associated with the reported clinical findings. Candidate regions were further interrogated using the National Genetic Database. Three fetuses from three distinct nuclear families in which the parents shared a similar ethnicity (either Ashkenazi or Bukharan Jews) but no reported consanguinity, were assessed. No copy number changes were observed, however evaluation of regions of homozygosity revealed a relevant candidate gene for the specific phenotype for each fetus. Using the National Genetic Database a specific mutation was examined in both parents (c.1167dupA mutation in the FKTN gene, c.167ins6[TTTCCC] mutation in the BSND gene and c.3761_3762delCCinsG in the PKHD1 gene, respectively). Both parents were found to be carriers and the fetuses homozygotes to the mutation. For the first time, specific sonographic pathology, data from ROH extracted from CMA results, and information regarding common founder mutations in distinct ethnic subgroups were combined to create a comprehensive streamlined approach to provide effective genetic diagnosis and counselling within the time constraints of ongoing pregnancy.

Estimating genotyping errors from genotype and reconstructed pedigree data

Abstract Genotyping errors are rules rather than exceptions in reality, and are found in virtually all but very small datasets. These errors, even when occurring at an extremely low rate, can derail many genetic analyses such as parentage/sibship assignments and linkage/association studies. Nonetheless, few robust and accurate methods are available for estimating the rate of occurrence of genotyping errors and for identifying individual erroneous genotypes at a locus. Methods based on duplicate genotyping are expensive, and estimate genotype inconsistency rather than error rate at a locus. Methods based on Hardy–Weinberg equilibrium tests have low robustness and low power, and apply only to those particular errors that cause excessive homozygosity. Methods based on pedigrees are powerful, robust and accurate. However, they rely on known and complete pedigrees that are unfortunately rarely available from natural populations in the wild. I proposed a maximum likelihood method to reconstruct pedigrees from genotype data with errors occurring at a roughly estimated (presumed) rate. In this paper, I describe how to use the method and inferred pedigree in estimating allelic dropout (or null allele) rate and false allele rate jointly at each marker locus, in identifying the erroneous genotypes, and in inferring the most likely genotypes at each locus of each individual. I examine the power, accuracy and robustness of the method by extensive simulations, and demonstrate the usefulness of the method by analysing three empirical datasets. It is concluded that, both pedigrees and the rates of genotyping errors at each locus can be reliably estimated from the same genotype data by the same likelihood method, when marker information is sufficient and some sampled individuals are first‐degree relatives. The erroneous genotypes are however inferred conservatively, and are reliably detected only when they occur in large families and/or at highly polymorphic loci. Estimation of genotyping error rates per locus and identification of erroneous genotypes of each individual at each locus should be routinely conducted to assess and improve data quality, to highlight markers for optimization of genotyping protocols or for replacement, and to enable the integration of genotyping errors in a robust statistical analysis.

HLA class I haplotype diversity is consistent with selection for frequent existing haplotypes.

The major histocompatibility complex (MHC) contains the most polymorphic genetic system in humans, the human leukocyte antigen (HLA) genes of the adaptive immune system. High allelic diversity in HLA is argued to be maintained by balancing selection, such as negative frequency-dependent selection or heterozygote advantage. Selective pressure against immune escape by pathogens can maintain appreciable frequencies of many different HLA alleles. The selection pressures operating on combinations of HLA alleles across loci, or haplotypes, have not been extensively evaluated since the high HLA polymorphism necessitates very large sample sizes, which have not been available until recently. We aimed to evaluate the effect of selection operating at the HLA haplotype level by analyzing HLA A~C~B~DRB1~DQB1 haplotype frequencies derived from over six million individuals genotyped by the National Marrow Donor Program registry. In contrast with alleles, HLA haplotype diversity patterns suggest purifying selection, as certain HLA allele combinations co-occur in high linkage disequilibrium. Linkage disequilibrium is positive (Dij'>0) among frequent haplotypes and negative (Dij'<0) among rare haplotypes. Fitting the haplotype frequency distribution to several population dynamics models, we found that the best fit was obtained when significant positive frequency-dependent selection (FDS) was incorporated. Finally, the Ewens-Watterson test of homozygosity showed excess homozygosity for 5-locus haplotypes within 23 US populations studied, with an average Fnd of 28.43. Haplotype diversity is most consistent with purifying selection for HLA Class I haplotypes (HLA-A, -B, -C), and was not inferred for HLA Class II haplotypes (-DRB1 and—DQB1). We discuss our empirical results in the context of evolutionary theory, exploring potential mechanisms of selection that maintain high linkage disequilibrium in MHC haplotype blocks.

Signatures of selection in mammalian clock genes with coding trinucleotide repeats: Implications for studying the genomics of high-pace adaptation.

Climate change is predicted to affect the reproductive ecology of wildlife; however, we have yet to understand if and how species can adapt to the rapid pace of change. Clock genes are functional genes likely critical for adaptation to shifting seasonal conditions through shifts in timing cues. Many of these genes contain coding trinucleotide repeats, which offer the potential for higher rates of change than single nucleotide polymorphisms (SNPs) at coding sites, and, thus, may translate to faster rates of adaptation in changing environments. We characterized repeats in 22 clock genes across all annotated mammal species and evaluated the potential for selection on repeat motifs in three clock genes (NR1D1,CLOCK, and PER1) in three congeneric species pairs with different latitudinal range limits: Canada lynx and bobcat (Lynx canadensis and L. rufus), northern and southern flying squirrels (Glaucomys sabrinus and G. volans), and white‐footed and deer mouse (Peromyscus leucopus and P. maniculatus). Signatures of positive selection were found in both the interspecific comparison of Canada lynx and bobcat, and intraspecific analyses in Canada lynx. Northern and southern flying squirrels showed differing frequencies at common CLOCK alleles and a signature of balancing selection. Regional excess homozygosity was found in the deer mouse at PER1 suggesting disruptive selection, and further analyses suggested balancing selection in the white‐footed mouse. These preliminary signatures of selection and the presence of trinucleotide repeats within many clock genes warrant further consideration of the importance of candidate gene motifs for adaptation to climate change.

New gSSR and EST-SSR markers reveal high genetic diversity in the invasive plant Ambrosia artemisiifolia L. and can be transferred to other invasive Ambrosia species.

Ambrosia artemisiifolia L., (common ragweed), is an annual invasive and highly troublesome plant species originating from North America that has become widespread across Europe. New sets of genomic and expressed sequence tag (EST) based simple sequence repeats (SSRs) markers were developed in this species using three approaches. After validation, 13 genomic SSRs and 13 EST-SSRs were retained and used to characterize the genetic diversity and population genetic structure of Ambrosia artemisiifolia populations from the native (North America) and invasive (Europe) ranges of the species. Analysing the mating system based on maternal families did not reveal any departure from complete allogamy and excess homozygosity was mostly due the presence of null alleles. High genetic diversity and patterns of genetic structure in Europe suggest two main introduction events followed by secondary colonization events. Cross-species transferability of the newly developed markers to other invasive species of the Ambrosia genus was assessed. Sixty-five percent and 75% of markers, respectively, were transferable from A. artemisiifolia to Ambrosia psilostachya and Ambrosia tenuifolia. 40% were transferable to Ambrosia trifida, this latter species being seemingly more phylogenetically distantly related to A. artemisiifolia than the former two.

Molecular Characterization of Mitochondrial D Loop of Indian Red Jungle Fowl

Mitochondrial DNA has the characteristic of quick evolution, matrilineal inheritance, and simple molecular structure, and it serves as the most used marker for molecular study. As an important role of genomics, studying it can help understand the origins, history, and adaptation of domestication. A 1325 bp fragment of the mtDNA D-loop region was amplified, sequenced and phylogenetic study was done. Out of 400 sites about 12 % were polymorphic between the Gallus gallus (G.g.) subspecies and most of them (60%) were transitions. Indian RJF showed very low genetic distances (0.008-0.013) with G.g. gallus birds from Thailand as compared to those from Japan and Indonesia. The G.g. murghi showed comparatively high genetic distance i.e. 031-0.034 with G.g. spadiceus and from 0.061-0.066 with G. gallus bankiva. The phylogenetic tree showed that G.g. banikva is well separated from the other three subspecies as it made a separate cluster. G.g. gallus makes two separate clusters i.e. one those from Thailand and other those from Japan and Indonesia. Within each chicken breed, there is an excess of homozygosity, but there is no significant reduction in the nucleotide diversity. Phenotypic modifications of chicken breeds as a result of artificial selection appear to stem from ancestral polymorphisms at a limited number of genetic loci.

P041 Next generation sequencing reveals HLA genomic and haplotype diversity in U.S. populations of European and African ancestry

Aim Next generation sequencing of extended long-range HLA gene products provides better molecular characterization of HLA genes and improves the assessment of HLA population diversity than traditional HLA genotyping methods. In addition, accurate comprehensive analyses of HLA diversity would be expected to increase the efficiency of unrelated donor selection. In this study we assessed the genomic and haplotype diversity in European Americans (EA) and African Americans (AFA). Methods We used a high-throughput NGS HLA typing method developed by Sirona Genomics to genotype class I (HLA-A, -B, -C) and class II (-DPA1, -DPB1, -DQA, -DQB, -DRB1, -DRB3, -DRB4, -DRB5) loci in >2400 EAs and >840 AFAs unrelated healthy individuals. Sample libraries were sequenced using Illumina HiSeq and NextSeq instruments. Sequence data was analyzed and genotypes were assigned using the MIA FORA NGS HLA software. Statistical analyses was performed using the GENE[RATE] computer pipeline (hla-net.eu). Results The distribution of allele frequencies in both EA and AFA populations is similar across all loci with an extreme right skew in each case. Rare alleles and novel DPA and DQB alleles were detected in both populations. The AFA population contained higher levels of mean heterozygosity at class I loci and most class II loci compared to the EA population. Tests of selective neutrality revealed a significant excess of homozygosity in B, C, and DPB1 in EAs and significant excess of heterozygosity in A and DQB1 in the AFA group. Haplotype analyses of neighbouring loci pairs in positive LD, such as B ∼ C and B ∼ DRB1, revealed novel associations at the 4th field level of resolution. Conclusions Our findings show unique 4-field associations that were not apparent in results obtained using testing methodologies with limited sequence coverage and phasing. In addition, our results provide new 4-field allele and haplotype frequencies. Our data provides a useful reference source for future population studies that in turn may allow for precise fine mapping of coding and non-coding sequences determining disease susceptibility and allo-immunogenicity. S. Krishnakumar: Employee; Company/Organization; Immucor. C. Wang: Employee; Company/Organization; Immucor. M. Li: Employee; Company/Organization; Immucor. M.N. Mindrinos: Employee; Company/Organization; Immucor.

ConStruct 1.0: An R Script to distinguish between substructure and consanguinity within a population using multilocus microsatellite data

ConStruct.1.r is an R Script that estimates the relative contributions of consanguinity and population substructure to excess homozygosity. ConStruct.1.r also offers the option of simulating data with a given FST and magnitude of consanguinity, incorporating a user-specified number of loci, number of alleles and population size. The method seems robust when population sizes are above 200 and individuals are genotyped at greater than 10 loci.

Likelihood Ratio Test for Excess Homozygosity at Marker Loci on X Chromosome.

The assumption of Hardy-Weinberg equilibrium (HWE) is generally required for association analysis using case-control design on autosomes; otherwise, the size may be inflated. There has been an increasing interest of exploring the association between diseases and markers on X chromosome and the effect of the departure from HWE on association analysis on X chromosome. Note that there are two hypotheses of interest regarding the X chromosome: (i) the frequencies of the same allele at a locus in males and females are equal and (ii) the inbreeding coefficient in females is zero (without excess homozygosity). Thus, excess homozygosity and significantly different minor allele frequencies between males and females are used to filter X-linked variants. There are two existing methods to test for (i) and (ii), respectively. However, their size and powers have not been studied yet. Further, there is no existing method to simultaneously detect both hypotheses till now. Therefore, in this article, we propose a novel likelihood ratio test for both (i) and (ii) on X chromosome. To further investigate the underlying reason why the null hypothesis is statistically rejected, we also develop two likelihood ratio tests for detecting (i) and (ii), respectively. Moreover, we explore the effect of population stratification on the proposed tests. From our simulation study, the size of the test for (i) is close to the nominal significance level. However, the size of the excess homozygosity test and the test for both (i) and (ii) is conservative. So, we propose parametric bootstrap techniques to evaluate their validity and performance. Simulation results show that the proposed methods with bootstrap techniques control the size well under the respective null hypothesis. Power comparison demonstrates that the methods with bootstrap techniques are more powerful than those without bootstrap procedure and the existing methods. The application of the proposed methods to a rheumatoid arthritis dataset indicates their utility.

Recurrent miscarriage is associated with the dopamine receptor (DRD2) genotype

The purpose of this research is to compare the prevalence of dopamine receptor D2 polymorphisms in women with recurrent miscarriage (RM) and healthy patients. Fifty-four women were enrolled in this case–control study. We performed DNA extraction of peripheral blood, followed by polymerase chain reaction to confirm single-strand polymorphisms and to sequence two polymorphisms: polymorphism 1 (rs6275) and polymorphism 2 (rs6277) in exon 7 of the dopamine receptor D2 (DRD2). The frequency of DRD2 polymorphism 2 (rs6277) was increased in the subjects with RM. An analysis of the DRD2 genotypes demonstrated an odds ratio of 2.37 (1.05–5.36, 95% confidence interval) for the polymorphism 2 (rs6277) in RM. The mean of the serum prolactin level was higher in the patients with RM (12.5 ng/ml) than in healthy women (8.1 ng/ml) p = 0.03. An excess homozygosity of the DRD2 polymorphism suggests a genetic predisposition to RMs, which could result in a mild serum prolactin increase. Thus, because of the potential role of prolactin in reproductive regulation, this polymorphism could play an important role in early pregnancy implantation and pregnancy maintenance.