Simultaneous fine mapping of closely linked epistatic quantitative trait loci using combined linkage disequilibrium and linkage with a general pedigree

Causal mutations and their intra- and inter-locus interactions play a critical role in complex trait variation. It is often not easy to detect epistatic quantitative trait loci (QTL) due to complicated population structure requirements for detecting epistatic effects in linkage analysis studies and due to main effects often being hidden by interaction effects. Mapping their positions is even harder when they are closely linked. The data structure requirement may be overcome when information on linkage disequilibrium is used. We present an approach using a mixed linear model nested in an empirical Bayesian approach, which simultaneously takes into account additive, dominance and epistatic effects due to multiple QTL. The covariance structure used in the mixed linear model is based on combined linkage disequilibrium and linkage information. In a simulation study where there are complex epistatic interactions between QTL, it is possible to simultaneously map interacting QTL into a small region using the proposed approach. The estimated variance components are accurate and less biased with the proposed approach compared with traditional models.     

Ignoring linkage disequilibrium among tightly linked markers induces false-positive evidence of linkage for affected sib pair analysis

Most multipoint linkage programs assume linkage equilibrium among the markers being studied. The assumption is appropriate for the study of sparsely spaced markers with intermarker distances exceeding a few centimorgans, because linkage equilibrium is expected over these intervals for almost all populations. However, with recent advancements in high-throughput genotyping technology, much denser markers are available, and linkage disequilibrium (LD) may exist among the markers. Applying linkage analyses that assume linkage equilibrium to dense markers may lead to bias. Here, we demonstrated that, when some or all of the parental genotypes are missing, assuming linkage equilibrium among tightly linked markers where strong LD exists can cause apparent oversharing of multipoint identity by descent (IBD) between sib pairs and false-positive evidence for multipoint model-free linkage analysis of affected sib pair data. LD can also mimic linkage between a disease locus and multiple tightly linked markers, thus causing false-positive evidence of linkage using parametric models, particularly when heterogeneity LOD score approaches are applied. Bias can be eliminated by inclusion of parental genotype data and can be reduced when additional unaffected siblings are included in the analysis.     

利用向日葵重组自交系构建遗传图谱

以向日葵自选系K55为母本、K58为父本杂交组合,通过单粒传得到的187个F_5：6代重组自交系群体为作图材料,联合应用SSR和AFLP标记构建遗传连锁图谱。经过78对SSR引物和48对AFLP引物组合选择性扩增,分别得到341和1119条带,共1460条,分别获得多态性条带184条和393条,共577条多态性条带,占所有条带的39.52%。SSR和AFLP标记各有84个和108个多态性标记偏离孟德尔分离比例(P=0.05),共192个偏分离标记。采用JoinMap4.0软件进行连锁分析,构建了1张总长度为2759.4 cM、包含17个连锁群、连锁495个多态性标记的遗传图谱,其中偏分离标记170个,标记间的平均图距为5.57 cM。每个连锁群上分布有5~72个标记,长68.88~250.17 cM。本图谱为向日葵永久性图谱,为向日葵重要性状QTL定位和基因克隆奠定基础。     

Assessment of parent-of-origin effects in linkage analysis of quantitative traits

Methods are presented for incorporation of parent-of-origin effects into linkage analysis of quantitative traits. The estimated proportion of marker alleles shared identical by descent is first partitioned into a component derived from the mother and a component derived from the father. These parent-specific estimates of allele sharing are used in variance-components or Haseman-Elston methods of linkage analysis so that the effect of the quantitative-trait locus carried on the maternally derived chromosome is potentially different from the effect of the locus on the paternally derived chromosome. Statistics for linkage between trait and marker loci derived from either or both parents are then calculated, as are statistics for testing whether the effect of the maternally derived locus is equal to that of the paternally derived locus. Analyses of data simulated for 956 siblings from 263 nuclear families who had participated in a linkage study revealed that type I error rates for these statistics were generally similar to nominal values. Power to detect an imprinted locus was substantially increased when analyzed with a model allowing for parent-of-origin effects, compared with analyses that assumed equal effects; for example, for an imprinted locus accounting for 30% of the phenotypic variance, the expected LOD score was 4.5 when parent-of-origin effects were incorporated into the analysis, compared with 3.1 when these effects were ignored. The ability to include parent-of-origin effects within linkage analysis of quantitative traits will facilitate genetic dissection of complex traits.     

A case study of the interpretation of linkage data using DNA fingerprinting probes

The statistical power of the technique of DNA fingerprinting relies greatly on the ability of the investigator to make an assumption that the presence or absence of different fingerprint bands are independent. Such linkage equilibrium is unlikely if bands are tightly linked to each other. Thus, when a new organism is to be investigated it is helpful to examine the segregation of parental bands into the offspring to confirm that the bands are not linked. By considering families of the tilapia Oreochrotnis niloticus , we produce statistical tests for linkage that can be applied to DNA fingerprint information. The use of these tests is more difficult and complicated if one parent is missing, but some progress can be made. We recommend that at least ten offspring are examined for segregation in families, since smaller numbers of offspring result in coincidental perfect agreements in band distributions in the absence of any linkage.     

Conclusion of LOD-score analysis for family data generated under two-locus models.

The power to detect linkage by the LOD-score method is investigated here for diseases that depend on the effects of two genes. The classical strategy is, first, to detect a major-gene (MG) effect by segregation analysis and, second, to seek for linkage with genetic markers by the LOD-score method using the MG parameters. We already showed that segregation analysis can lead to evidence for a MG effect for many two-locus models, with the estimates of the MG parameters being very different from those of the two genes involved in the disease. We show here that use of these MG parameter estimates in the LOD-score analysis may lead to a failure to detect linkage for some two-locus models. For these models, use of the sib-pair method gives a non-negligible increase of power to detect linkage. The linkage-homogeneity test among subsamples differing for the familial disease distribution provides evidence of parameter misspecification, when the MG parameters are used. Moreover, for most of the models, use of the MG parameters in LOD-score analysis leads to a large bias in estimation of the recombination fraction and sometimes also to a rejection of linkage for the true recombination fraction. A final important point is that a strong evidence of an MG effect, obtained by segregation analysis, does not necessarily imply that linkage will be detected for at least one of the two genes, even with the true parameters and with a close informative marker.     

The role of the sulfonium linkage in the stabilization of the ferrous form of myeloperoxidase: a comparison with lactoperoxidase

In all mammalian peroxidases, the heme is covalently attached to the protein via two ester linkages between conserved aspartate (Asp94) and glutamate residues (Glu242) and modified methyl groups on pyrrole rings A and C. Only myeloperoxidase has an additional sulfonium ion linkage between the sulfur atom of the conserved methionine 243 and the beta-carbon of the vinyl group on pyrrole ring A. Upon reduction from Fe(III) to Fe(II), lactoperoxidase (LPO) but not myeloperoxidase (MPO) is shown to adopt three distinct active site conformations which depend on pH and time. Comparative spectroscopic analysis (UV-Vis absorption and resonance Raman) of the ferrous forms of LPO, wild-type MPO and the variants Asp94Val, Glu242Gln, Met243Thr and Met243Val clearly demonstrate that a single, stable ferrous form of MPO is present only in those proteins which retain an intact sulfonium linkage. By contrast, both ferrous Met243Thr and Met243Val can assume two conformations. They resemble ferrous LPO, being five-coordinated high-spin species that are distinguished by the strength of the proximal Fe-histidine bond. This bond weakens with time or decreasing pH, as indicated by the Fe-histidine stretching bands.     

On the use of DNA fingerprints for linkage studies in cattle

To find a marker for the bovine "muscular hypertrophy" gene and for the "roan" locus, we have typed six cattle pedigrees totaling 540 animals for nine blood group systems, for 12 biochemical markers, for RFLPs at four loci, and with five probes revealing multilocus DNA fingerprints. Segregation analysis of the fingerprint bands showed that, in cattle, a fingerprint probe will reveal a mean of 7.6 clearly resolvable bands, behaving as simple, highly informative Mendelian entities characterized by a mean mutation rate of +/- 1/4500 gametes. For one of the bands, we observed a "mutation burst" generating germline mosaicism. Because some of the fingerprint bands were allelic or corresponded to clustered minisatellites, a mean of only 5.7 independent loci is explored per probe. Fingerprint bands revealed by different probes also show a clear propensity for close linkage, pointing toward nonrandom distribution of minisatellite sequences or the existence of minisatellite clusters. Although this reduces the power of fingerprints for linkage analysis substantially, we were able to demonstrate genetic linkage between fingerprint bands and at least three of the classical markers, to exclude the roan locus from 4.5 Morgans of the bovine genome with the DNA fingerprints and for an additional 2.5 Morgans with the classical markers, and to identify a solid candidate marker for the bovine muscular hypertrophy gene, yielding a lod score greater than or equal to 2.84 without any obliged recombinant.     

Gene Flow and Selection in a Two-Locus System

A model of gene flow and selection in two linked loci is analyzed. The problems considered are the effects of linkage on the clines in frequencies at the two loci and the role of gene flow in producing linkage disequilibrium between the loci. Also, the possible significance of linkage as a mechanism for permitting a population of "track" spatial changes in the environment is considered. The results are that when the recombination fraction between the loci is of the same order of magnitude as the selection coefficients or smaller, then linkage is important in determining the gene frequencies and a substantial amount of linkage disequilibrium is present in the cline. Depending on the spatial pattern of selection on the two loci, linkage can either decrease or increase a population's response to local selection.     

The effects of deleterious mutations on linked, neutral variation in small populations.

The effects of recessive, deleterious mutations on genetic variation at linked neutral loci can be heterozygosity-decreasing because of reduced effective population sizes or heterozygosity-increasing because of associative overdominance. Here we examine the balance between these effects by simulating individual diploid genotypes in small panmictic populations. The haploid genome consists of one linkage group with 1000 loci that can have deleterious mutations and a neutral marker. Combinations of the following parameters are studied: gametic mutation rate to harmful alleles (U), population size (N), recombination rate (r), selection coefficient (s), and dominance (h). Tight linkage (r 相似文献   

Tyrosine sulfation site is located in the C-terminal fibrin-binding domain in secreted fibronectin from rat embryo fibroblasts, line 3Y1

Tryptic fragments of [35S]sulfate-labeled 3Y1 secreted fibronectin were fractionated by hydroxylapatite column chromatography and examined using sodium dodecyl sulfate gel electrophoresis, followed by autoradiography. Radioactive bands containing tyrosine-O-[35S]sulfate were detected at 17- and 40-kDa positions under reducing conditions. Under nonreducing conditions, the 17-kDa band was no longer present and new bands at 57- and 80-kDa positions appeared, indicating a disulfide linkage between the two smaller fragments in the native state. These fragments exhibited binding affinity toward fibrin and could be immunoprecipitated by the monoclonal antifibronectin Fib-2 domain antibody. These results suggested that the tyrosine sulfation site in 3Y1 secreted fibronectin is located in the C-terminal fibrin-binding (Fib-2) domain, being within 17 kDa of the C-terminus.     

Keep Your Opponents Close: Social Context Affects EEG and fEMG Linkage in a Turn-Based Computer Game

In daily life, we often copy the gestures and expressions of those we communicate with, but recent evidence shows that such mimicry has a physiological counterpart: interaction elicits linkage, which is a concordance between the biological signals of those involved. To find out how the type of social interaction affects linkage, pairs of participants played a turn-based computer game in which the level of competition was systematically varied between cooperation and competition. Linkage in the beta and gamma frequency bands was observed in the EEG, especially when the participants played directly against each other. Emotional expression, measured using facial EMG, reflected this pattern, with the most competitive condition showing enhanced linkage over the facial muscle-regions involved in smiling. These effects were found to be related to self-reported social presence: linkage in positive emotional expression was associated with self-reported shared negative feelings. The observed effects confirmed the hypothesis that the social context affected the degree to which participants had similar reactions to their environment and consequently showed similar patterns of brain activity. We discuss the functional resemblance between linkage, as an indicator of a shared physiology and affect, and the well-known mirror neuron system, and how they relate to social functions like empathy.     

Ignoring intermarker linkage disequilibrium induces false-positive evidence of linkage for consanguineous pedigrees when genotype data is missing for any pedigree member

Missing genotype data can increase false-positive evidence for linkage when either parametric or nonparametric analysis is carried out ignoring intermarker linkage disequilibrium (LD). Previously it was demonstrated by Huang et al. [1] that no bias occurs in this situation for affected sib-pairs with unrelated parents when either both parents are genotyped or genotype data is available for two additional unaffected siblings when parental genotypes are missing. However, this is not the case for autosomal recessive consanguineous pedigrees, where missing genotype data for any pedigree member within a consanguinity loop can increase false-positive evidence of linkage. False-positive evidence for linkage is further increased when cryptic consanguinity is present. The amount of false-positive evidence for linkage, and which family members aid in its reduction, is highly dependent on which family members are genotyped. When parental genotype data is available, the false-positive evidence for linkage is usually not as strong as when parental genotype data is unavailable. For a pedigree with an affected proband whose first-cousin parents have been genotyped, further reduction in the false-positive evidence of linkage can be obtained by including genotype data from additional affected siblings of the proband or genotype data from the proband's sibling-grandparents. For the situation, when parental genotypes are unavailable, false-positive evidence for linkage can be reduced by including genotype data from either unaffected siblings of the proband or the proband's married-in-grandparents in the analysis.     

Measurement of the screw-home motion of the knee is sensitive to errors in axis alignment

Measurements of joint angles during motion analysis are subject to error caused by kinematic crosstalk, that is, one joint rotation (e. g., flexion) being interpreted as another (e.g., abduction). Kinematic crosstalk results from the chosen joint coordinate system being misaligned with the axes about which rotations are assumed to occur. The aim of this paper is to demonstrate that measurement of the so-called "screw-home" motion of the human knee, in which axial rotation and extension are coupled, is especially prone to errors due to crosstalk. The motions of two different two-segment mechanical linkages were examined to study the effects of crosstalk. The segments of the first linkage (NSH) were connected by a revolute joint, but the second linkage (SH) incorporated gearing that caused 15 degrees of screw-home rotation to occur with 90 degrees knee flexion. It was found that rotating the flexion axis (inducing crosstalk) could make linkage NSH appear to exhibit a screw-home motion and that a different rotation of the flexion axis could make linkage SH apparently exhibit pure flexion. These findings suggest that the measurement of screw-home rotation may be strongly influenced by errors in the location of the flexion axis. The magnitudes of these displacements of the flexion axis were consistent with the inter-observer variability seen when five experienced observers defined the flexion axis by palpating the medial and lateral femoral epicondyles. Care should be taken when interpreting small internal-external rotations and abduction-adduction angles to ensure that they are not the products of kinematic crosstalk.     

A cladistic analysis of phenotypic associations with haplotypes inferred from restriction endonuclease mapping. I. Basic theory and an analysis of alcohol dehydrogenase activity in Drosophila

Because some genes have been cloned that have a known biochemical or physiological function, genetic variation can be measured in a population at loci that may directly influence a phenotype of interest. With this measured genotype approach, specific alleles or haplotypes in the probed DNA region can be assigned phenotypic effects. In this paper we address several problems encountered in implementing the measured genotype approach with restriction site data. A number of analytical problems arise in part as a consequence of the linkage disequilibrium that is commonly encountered when dealing with small DNA regions: 1) different restriction site polymorphisms are not statistically independent, 2) the sites being measured are not likely to be the direct cause of the associated phenotypic effects, 3) haplotype classes may be phenotypically heterogeneous, and 4) the sites that are most strongly associated with phenotypic effects are not necessarily the most closely linked to the actual genetic cause of the effects. When recombination and gene conversion are rare, the primary cause of linkage disequilibrium is history (mutational origin, genetic drift, hitchhiking, etc.). We deal with historical association directly by producing a cladogram that partially reconstructs the evolutionary history of the present-day haplotype variability. The cladogram defines a nested analysis of variance that simultaneously detects phenotypic effects, localizes the effects within the cladogram, and identifies haplotypes that are potentially heterogeneous in their phenotypic associations. The power of this approach is illustrated by an analysis of the associations between alcohol dehydrogenase (ADH) activity and restriction site variability in a 13-kb fragment surrounding the ADH locus in Drosophila melanogaster.     

RFLP linkage map of asparagus.

A population resulting from a double pseudotestcross of two outbred-derived asparagus (Asparagus officinalis L.) clones was evaluated by RFLP (restriction fragment length polymorphism) analysis to produce individual maps of the male and female parents. An asparagus PstI genomic library was created and used as the source of probes. Scoring of bands was done by examining SDRFs (single dose restriction fragments) that are present in one parent and absent in the other and segregate 1:1 in the progeny. The data were analyzed as a backcross population; inversion or recoding allowed for the detection of repulsion phase linkage. The male parent map consisted of 33 loci in 10 groups, while the female parent map had 48 loci arranged in 14 groups. Segregation distortion was minimal (5%), and 17% of the markers were found to be unlinked. Loci of the configuration a/b x a/b and a/c x b/c were used to bridge seven homologous linkage groups between the two parents. The sex locus was not found to be associated with any linkage group. Key words : RFLP, bridge loci, repulsion phase linkage, double pseudotestcross.     

Accounting for linkage in family-based tests of association with missing parental genotypes

In studies of complex diseases, a common paradigm is to conduct association analysis at markers in regions identified by linkage analysis, to attempt to narrow the region of interest. Family-based tests for association based on parental transmissions to affected offspring are often used in fine-mapping studies. However, for diseases with late onset, parental genotypes are often missing. Without parental genotypes, family-based tests either compare allele frequencies in affected individuals with those in their unaffected siblings or use siblings to infer missing parental genotypes. An example of the latter approach is the score test implemented in the computer program TRANSMIT. The inference of missing parental genotypes in TRANSMIT assumes that transmissions from parents to affected siblings are independent, which is appropriate when there is no linkage. However, using computer simulations, we show that, when the marker and disease locus are linked and the data set consists of families with multiple affected siblings, this assumption leads to a bias in the score statistic under the null hypothesis of no association between the marker and disease alleles. This bias leads to an inflated type I error rate for the score test in regions of linkage. We present a novel test for association in the presence of linkage (APL) that correctly infers missing parental genotypes in regions of linkage by estimating identity-by-descent parameters, to adjust for correlation between parental transmissions to affected siblings. In simulated data, we demonstrate the validity of the APL test under the null hypothesis of no association and show that the test can be more powerful than the pedigree disequilibrium test and family-based association test. As an example, we compare the performance of the tests in a candidate-gene study in families with Parkinson disease.     

Mapping of a gene for familial juvenile nephronophthisis: Refining the map and defining flanking markers on chromosome 2

Familial juvenile nephronophthisis (NPH) is an autosomal recessive kidney disease that leads to end-stage renal failure in adolescence and is associated with the formation of cysts at the cortico-medullary junction of the kidneys. NPH is responsible for about 15% of end-stage renal disease in children, as shown by Kleinknecht and Habib. NPH in combination with autosomal recessive retinitis pigmentosa is known as the Senior-Løken syndrome (SLS) and exhibits renal pathology that is identical to NPH. We had excluded 40% of the human genome from linkage with a disease locus for NPH or SLS when antignac et al. first demonstrated linkage for an NPH locus on chromosome 2. We present confirmation of linkage of an NPH locus to microsatellite markers on chromosome 2 in nine families with NPH. By linkage analysis with marker AFM262xb5 at locus D2S176, a maximum lod score of 5.05 at a θ_max = .03 was obtained. In a large NPH family that yielded at D2S176 a maximum lod score of 2.66 at θ_max = .0, markers AFM172xc3 and AFM016yc5, representing loci D2S135 and D2S110, respectively, were identified as flanking markers, thereby defining the interval for an NPH locus to a region of approximately 15 cM. Furthermore, the cytogenetic assignment of the NPH region was specified to 2p12-(2q13 or adjacent bands) by calculation of linkage between these flanking markers and markers with known unique cytogenetic assignment. The refined map may serve as a genetic framework for additional genetic and physical mapping of the region.     

Linkage analysis of quantitative trait loci in the presence of heterogeneity

Variance component modeling for linkage analysis of quantitative traits is a powerful tool for detecting and locating genes affecting a trait of interest, but the presence of genetic heterogeneity will decrease the power of a linkage study and may even give biased estimates of the location of the quantitative trait loci. Many complex diseases are believed to be influenced by multiple genes and therefore genetic heterogeneity is likely to be present for many real applications of linkage analysis. We consider a mixture of multivariate normals to model locus heterogeneity by allowing only a proportion of the sampled pedigrees to segregate trait-influencing allele(s) at a specific locus. However, for mixtures of normals the classical asymptotic distribution theory of the maximum likelihood estimates does not hold, so tests of linkage and/or heterogeneity are evaluated using resampling methods. It is shown that allowing for genetic heterogeneity leads to an increase in power to detect linkage. This increase is more prominent when the genetic effect of the locus is small or when the percentage of pedigrees not segregating trait-influencing allele(s) at the locus is high.     

The transmission/disequilibrium test: history, subdivision, and admixture.

Disease association with a genetic marker is often taken as a preliminary indication of linkage with disease susceptibility. However, population subdivision and admixture may lead to disease association even in the absence of linkage. In a previous paper, we described a test for linkage (and linkage disequilibrium) between a genetic marker and disease susceptibility; linkage is detected by this test only if association is also present. This transmission/disequilibrium test (TDT) is carried out with data on transmission of marker alleles from parents heterozygous for the marker to affected offspring. The TDT is a valid test for linkage and association, even when the association is caused by population subdivision and admixture. In the previous paper, we did not explicitly consider the effect of recent history on population structure. Here we extend the previous results by examining in detail the effects of subdivision and admixture, viewed as processes in population history. We describe two models for these processes. For both models, we analyze the properties of (a) the TDT as a test for linkage (and association) between marker and disease and (b) the conventional contingency statistic used with family data to test for population association. We show that the contingency test statistic does not have a chi 2 distribution if subdivision or admixture is present. In contrast, the TDT remains a valid chi 2 statistic for the linkage hypothesis, regardless of population history.