DSLINK: a computer program for gene-centromere linkage analysis in families with a trisomic offspring.

Trisomic individuals provide information for gene-centromere mapping, since two of the four chromatids in a meiotic tetrad can be recovered. When centromeric markers are available, linkage analysis between the centromere and any marker locus can be performed in nuclear families having one or more trisomic offspring. Since conventional linkage programs consider only disomic individuals, we have written a FORTRAN computer program, DSLINK, that performs gene-centromere linkage analysis on the basis of information on trisomic and disomic offspring. This program makes it possible to study the relationship between recombination and chromosome segregation.     

Genetics and biology of human ovarian teratomas. II. Molecular analysis of origin of nondisjunction and gene-centromere mapping of chromosome I markers.

Chromosomal heteromorphisms and DNA polymorphisms have been utilized to identify the mechanisms that lead to formation of human ovarian teratomas and to construct a gene-centromere map of chromosome 1 by using those teratomas that arise by meiotic nondisjunction. Of 61 genetically informative ovarian teratomas, 21.3% arose by nondisjunction at meiosis I, and 39.3% arose by meiosis II nondisjunction. Eight polymorphic marker loci on chromosome 1p and one marker on 1q were used to estimate a gene-centromere map. The results show clear linkage of the most proximal 1p marker (NRAS) and the most proximal 1q marker (D1S61) to the centromere at a distance of 14 cM and 20 cM, respectively. Estimated gene-centromere distances suggest that, while recombination occurs normally in ovarian teratomas arising by meiosis II errors, ovarian teratomas arising by meiosis I nondisjunction have altered patterns of recombination. Furthermore, the estimated map demonstrates clear evidence of chiasma interference. Our results suggest that ovarian teratomas can provide a rapid method for mapping genes relative to the centromere.     

Improving linkage analysis in outcrossed forest trees - an example from Acacia mangium

Mapping in forest trees generally relies on outbred pedigrees in which genetic segregation is the result of meiotic recombination from both parents. The currently available mapping packages are not optimal for outcrossed pedigrees as they either cannot order phase-ambiguous data or only use pairwise information when ordering loci within linkage groups. A new package, OUTMAP, has been developed for mapping codominant loci in outcrossed trees. A comparison of maps produced using linkage data from two pedigrees of Acacia mangium Willd demonstrated that the marker orders produced using OUTMAP were consistently of higher likelihood than those produced by JOINMAP. In addition, the maps were produced more efficiently, without the need for recoding data or the detailed investigation of pairwise recombination fractions which was necessary to select the optimal marker order using JOINMAP. Distances between markers often varied from those calculated by JOINMAP, resulting in an increase in the estimated genome length. OUTMAP can be used with all segregation types to determine phase and to calculate the likelihood of alternative marker orders, with a choice of three optimisation methods.     

Methods for studying recombination on chromosomes that undergo nondisjunction

A lod score method is provided for mapping genes relative to the centromere using family data from autosomal trisomies. Such gene-centromere mapping can be performed whenever two or more members of a meiotic tetrad can be recovered. The critical mapping parameter is not the recombination value theta or the map distance omega, but the probability of nonreduction in a heterozygous host, the probability of heterozygosity (nonreduction) is 1-gamma/2 for a meiosis I error and gamma for a meiosis II error. Under various assumptions regarding chiasma interference, gamma can be related to theta and omega. We provide specific methods for estimating gamma and theta from trisomy data using maximum likelihood, so that recombination may be studied on chromosomes that underwent nondisjunction.     

Estimation of gametic frequencies from F2 populations using the EM algorithm and its application in the analysis of crossover interference in rice

The gametes produced in meiosis provide information on the frequency of recombination and also on the interdependence of recombination events, i.e. interference. Using F₂ individuals, it is not possible in all cases to derive the gametes, which have fused, and which provide the information about interference unequivocally when three or more segregating markers are considered simultaneously. Therefore, a method was developed to estimate the gametic frequencies using a maximum likelihood approach together with the expectation maximisation algorithm. This estimation procedure was applied to F₂ mapping data from rice (Oryza sativa L.) to carry out a genome-wide analysis of crossover interference. The distribution of the coefficient of coincidence in dependence on the recombination fraction revealed for all chromosomes increasing positive interference with decreasing interval size. For some chromosomes this mutual inhibition of recombination was not so strong in small intervals. The centromere had a significant effect on interference. The positive interference found in the chromosome arms were reduced significantly when the intervals considered spanned the centromere. Two chromosomes even demonstrated independent recombination and slightly negative interference for small intervals including the centromere. Different marker densities had no effect on the results. In general, interference depended on the frequency of recombination events in relation to the physical length. The strength of the centromere effect on interference seemed to depend on the strength of recombination suppression around the centromere.Electronic Supplementary Material Supplementary material is available for this article at     

Influence of genetic background and heterozygosity on meiotic recombination in Arabidopsis thaliana.

Plant breeding relies on genetic variability generated by meiotic recombination. Control of recombination frequencies is not yet possible, but would significantly extend the options for plant-breeding strategies. A prerequisite would be variability of recombination frequencies. In this study, 15 transgenic kanamycin (KR) and hygromycin (HR) resistance gene insertions mapping to the five Arabidopsis thaliana chromosomes were used as genetic markers. Recombination frequencies were determined from the frequencies of resistance phenotypes within populations segregating for linked KR and HR markers. Recombination frequencies of marker pairs were compared among these four ecotypes, among F1s in both reciprocal forms derived from these ecotypes, and between F1s and their parent lines. On average, the recombination frequencies in F1 crosses were substantially higher (up to 2-fold) than in the homozygous parental ecotypes. A strong negative correlation between genetic similarities of ecotypes and recombination frequencies was detected for two adjacent marker pairs located on the long arm of chromosome 3, but not for marker pairs in other genomic regions. Our results suggest that heterozygosity influences recombination in plant breeding, and cannot be ignored in genetic mapping of genomes.     

Fast and Accurate Construction of Ultra-Dense Consensus Genetic Maps Using Evolution Strategy Optimization

Our aim was to develop a fast and accurate algorithm for constructing consensus genetic maps for chip-based SNP genotyping data with a high proportion of shared markers between mapping populations. Chip-based genotyping of SNP markers allows producing high-density genetic maps with a relatively standardized set of marker loci for different mapping populations. The availability of a standard high-throughput mapping platform simplifies consensus analysis by ignoring unique markers at the stage of consensus mapping thereby reducing mathematical complicity of the problem and in turn analyzing bigger size mapping data using global optimization criteria instead of local ones. Our three-phase analytical scheme includes automatic selection of ~100-300 of the most informative (resolvable by recombination) markers per linkage group, building a stable skeletal marker order for each data set and its verification using jackknife re-sampling, and consensus mapping analysis based on global optimization criterion. A novel Evolution Strategy optimization algorithm with a global optimization criterion presented in this paper is able to generate high quality, ultra-dense consensus maps, with many thousands of markers per genome. This algorithm utilizes "potentially good orders" in the initial solution and in the new mutation procedures that generate trial solutions, enabling to obtain a consensus order in reasonable time. The developed algorithm, tested on a wide range of simulated data and real world data (Arabidopsis), outperformed two tested state-of-the-art algorithms by mapping accuracy and computation time.     

Application of the Ovarian Teratoma Mapping Method in the Mouse

Murine ovarian teratomas were used to determine recombination percentages for gene-gene and centromere-gene intervals. Data were obtained utilizing a recombinant inbred strain, LTXBJ, and a number of newly developed LT/SvEi congenic strains.--Centromere-gene recombination was measured at 11.3 +/- 1.2% for the centromere of chromosome 7 - Gpi-1 interval and 15.8 +/- 2.4% for the centromere of chromosome 14 - Np-1 interval using the ovarian teratoma method. The centromere - Np-1 interval was measured at 26.5 +/- 3.6% using a standard backcross involving the Rb6Bnr Robertsonian translocation as a centromere marker.--To assess the accuracy of the ovarian teratoma mapping method, we compared the recombination frequency obtained for the Mpi-1-Mod-1 interval on chromosome 9 using the ovarian teratoma method to that obtained using a standard backcross. The recombination percentage was 22.9 +/- 5.4 using the ovarian teratoma method and 18.6 +/- 3.3 using the backcross method, indicating that the two methods produce equivalent estimates of recombination. In addition, for centromere-gene intervals known to be more than 30 cM in length, the ovarian teratoma method was consistent with classical recombination methods, yielding high recombination percentages. We conclude from these results that the ovarian teratoma mapping method is a reliable method for estimating recombination frequencies and the most accurate method available for estimating centromere-gene recombination frequency in the mouse.     

Comparative gene mapping in Arabidopsis lyrata chromosomes 6 and 7 and A. thaliana chromosome IV: evolutionary history, rearrangements and local recombination rates

We have increased the density of genetic markers on the Arabidopsis lyrata chromosomes AL6 and AL7 corresponding to the A. thaliana chromosome IV, in order to determine chromosome rearrangements between these two species, and to compare recombination fractions across the same intervals. We confirm the two rearrangements previously inferred (a reciprocal translocation and a large inversion, which we infer to be pericentric). By including markers around the centromere regions of A. thaliana chromosomes IV and V, we localize the AL6 centromere, and can localize the breakpoints of these chromosome rearrangements more precisely than previously. One translocation breakpoint was close to the centromere, and the other coincided with one end of the inversion, suggesting that a single event caused both rearrangements. At the resolution of our mapping, apart from these rearrangements, all other markers are in the same order in A. lyrata and A. thaliana. We could thus compare recombination rates in the two species. We found slightly higher values in A. thaliana, and a minimum estimate for regions not close to a centromere in A. lyrata is 4-5 centimorgans per megabase. The mapped region of AL7 includes the self-incompatibility loci (S-loci), and this region has been predicted to have lower recombination than elsewhere in the genome. We mapped 17 markers in a region of 1.23 Mb surrounding these loci, and compared the approximately 600 kb closest to the S-loci with the surrounding region of approximately the same size. There were significantly fewer recombination events in the closer than the more distant region, supporting the above prediction, but showing that the low recombination region is very limited in size.     

Construction of an AFLP genetic map with nearly complete genome coverage in Pinus taeda

  De novo construction of complete genetic linkage maps requires large mapping populations, large numbers of genetic markers, and efficient algorithms for ordering markers and evaluating order confidence. We constructed a complete genetic map of an individual loblolly pine (Pinus taeda L.) using amplified fragment length polymorphism (AFLP) markers segregating in haploid megagametophytes and PGRI mapping software. We generated 521 polymorphic fragments from 21 AFLP primer pairs. A total of 508 fragments mapped to 12 linkage groups, which is equal to the Pinus haploid chromosome number. Bootstrap locus order matrices and recombination matrices generated by PGRI were used to select 184 framework markers that could be ordered confidently. Order support was also evaluated using log likelihood criteria in MAPMAKER. Optimal marker orders from PGRI and MAPMAKER were identical, but the implied reliability of orders differed greatly. The framework map provides nearly complete coverage of the genome, estimated at approximately 1700 cM in length using a modified estimator. This map should provide a useful framework for merging existing loblolly pine maps and adding multiallelic markers as they become available. Map coverage with dominant markers in both linkage phases will make the map useful for subsequent quantitative trait locus mapping in families derived by self-pollination. Received: 7 August 1998 / Accepted: 27 October 1998     

Point and interval estimates of marker location in radiation hybrid mapping.

Radiation hybrid (RH) mapping is a powerful method for ordering loci on chromosomes and for estimating the distances between them. RH mapping is currently used to construct both framework maps, in which all markers are ordered with high confidence (e.g., 1,000:1 relative maximum likelihood), and comprehensive maps, which include markers with less-confident placement. To deal with uncertainty in the order and location of markers, marker positions may be estimated conditional on the most likely marker order, plausible intervals for nonframework markers may be indicated on a framework map, or bins of markers may be constructed. We propose a statistical method for estimating marker position that combines information from all plausible marker orders, gives a measure of uncertainty in location for each marker, and provides an alternative to the current practice of binning. Assuming that the prior distribution for the retention probabilities is uniform and that the marker loci are distributed independently and uniformly on an interval of specified length, we calculate the posterior distribution of marker position for each marker. The median or mean of this distribution provides a point estimate of marker location. An interval estimate of marker location may be constructed either by using the 100(alpha/2) and 100(1-alpha)/2 percentiles of the distribution to form a 100(1-alpha) % posterior credible interval or by calculating the shortest 100(1-alpha) % posterior credible interval. These point and interval estimates take into account ordering uncertainty and do not depend on the assumption of a particular marker order. We evaluate the performance of the estimates on the basis of results from simulated data and illustrate the method with two examples.     

三点自交法和两点最大似然法构建水稻F2群体分子标记连锁图谱的比较

根据连锁遗传的原理,列出了三点自交法和两点自交最大似然(ML)法估算显性标记遗传距离的具体步骤和算法,将水稻F2群体含香味基因Aro及其连锁的RFLP数据转变为显性标记数据后,用上述两种方法构建的连锁图谱与用MAPMAKER软件计算共显性数据得到的图谱排序相同、标记间距离相近．但是标记数据存在较大程度偏分离时,由三点自交法构建的图谱中标记间图距有增大趋势．作者为提高作图精确性,简化计算过程,讨论了三点自交法对估算重组值的影响及其在分子标记作图中的应用价值,并建议将共显性标记转变为显性标记时进行两次自交ML法估算。     

Pollen markers for gene-centromere mapping in diploid potato

The utility of two pollen genetic markers for estimating the extent of meiotic recombination between the centromere and a marker gene was tested in 2n pollen of diploid potato clones. One of these markers was the distal locus amylose-free (amf) on chromosome 8 and the other was the isozyme locus alcohol dehydrogenase (Adh-1) on chromosome 4. In the case of the amf locus, the gene-centromere distance was estimated in a normal synaptic and a desynaptic genotype. In both cases the genetic analysis was confined to: (1) a direct estimation of the phenotypic (blue vs red) segregation ratios in FDR (first-division restitution) 2n pollen and (2) a classification of the 4 x progeny from 4x (nulliplex amf) x 2x (Amf/amf) crosses into duplex, simplex and nulliplex classes. The recombination frequency between the centromere and the amf locus in the normal synaptic genotype B92-7015-4 corresponded to a gene-centromere distance of 48.8 cM, whereas this distance amounted to 13.3 cM in the desynaptic genotype RS93-8025-1. Hence desynapsis reduced crossing-over by 73%. The observed genetic distance of 48.8 cM in the normal synaptic clone, B92-7015-4, is the highest gene-centromere distance reported so far in potato and this could be explained on the assumption of absolute chiasma interference. For the Adh-1 locus, it was found that heterozygous 2n pollen grains could be detected in pollen samples of the diploid clones, because of the occurrence of a heterodimeric band of the isozyme. Unlike the amf locus, the genecentromere distance for the Adh-1 locus was estimated only on the basis of the duplex, simplex and nulliplex classes in the progenies from 4x (nulliplex Adh-1 ² )x B92-7015-4 (Adh-1 ¹ /Adh-1 ² )crosses and was found to be 19.4 cM. Because the accurate positions of centromeres in relation to other loci are not available in the existing genetic maps of potato, which are saturated with molecular markers, halftetrad analysis is a promising additional approach to the basic genetics of this crop.     

Linkage analysis for mapping genes of sex-influenced traits

Abstract Statistical methods are developed to estimate gender-specific and gender-average recombination frequencies between a biallelic or multiallelic marker and a sex-influenced gene. Iterative solutions are developed for intercross (or F-2 design). For biallelic markers, two iterative solutions are required, one for coupling and repulsion parental linkage phases and one for mixed parental linkage phases. For multiallelic markers, one set of iterative equations applies to all possible parental linkage phases. The resulting formulae for estimating recombination frequency use the full data set and yield estimates that are exactly the same as the true parameters if the observed and expected phenotypic distributions are equal. When one parent is homozygous for the sex-influenced gene as is expected with the backcross design, simple solutions exist for estimating recombination frequencies. However, offspring of one gender (male or female) do not have linkage information depending on whether the homozygous parent has two male-dominant or male-recessive alleles. Consequently, an F-2 design is more efficient than a backcross design for mapping a sex-influenced gene. Knowing each parental linkage phase is important to apply the methods developed in this article. It is shown that an individual's linkage phase of the sex-influenced locus can be determined based on allele transmission from the parents for all crosses except under the mating between an expressed male and an unexpressed female.     

Mapping disease genes: family-based association studies.

With recent rapid advances in mapping of the human genome, including highly polymorphic and closely linked markers, studies of marker associations with disease are increasingly relevant for mapping disease genes. The use of nuclear-family data in association studies was initially developed to avoid possible ethnic mismatching between patients and randomly ascertained controls. The parental marker alleles not transmitted to an affected child or never transmitted to an affected sib pair form the so-called AFBAC (affected family-based controls) population. In this paper, the theoretical foundation of the AFBAC method is proved for any single-locus model of disease and for any nuclear family-based ascertainment scheme. In a random-mating population, when there is a marker association with disease, the AFBAC population provides an unbiased estimate of the overall population (control) marker alleles when the recombination fraction (theta) between the marker and disease genes is sufficiently small that it can be taken as zero (theta = 0). With population stratification, only marker associations present in the subpopulations will be detected with family-based analyses. Of more importance, however, is the fact that, when theta not equal to 0, differences between transmitted parental (patient) marker allele frequencies and non- or never-transmitted parental marker allele frequencies (implying a marker association with disease) can only be observed for marker genes linked to a disease gene (theta < 1/2). Thus, associations of unlinked marker loci with disease at the population level, caused by population stratification, migration, or admixture, are eliminated. This validates the use of family-based association tests as an appropriate strategy for mapping disease genes.     

Microsatellite-centromere mapping in the loach, Misgurnus anguillicaudatus

Primer sets for 15 polymorphic microsatellite loci were developed in the loach, Misgurnus anguillicaudatus (Cobitidae) by molecular cloning and sequencing techniques. Mendelian inheritance was confirmed for the 15 loci by examining the genotypic segregation produced with the primer sets in two full-sib families. The loci were mapped in relation to their centromere in four gynogenetic diploid lines, which were induced by inhibition of the second meiotic division after fertilization with genetically inert sperm. Microsatellite-centromere recombination rates ranged between 0.06 and 0.95 under the assumption of complete interference. Thus, these loci are distributed from the centromeres to the telomeres of their respective chromosomes. The success of mitotic gynogenesis, produced by suppression of the first cleavage, was verified by homozygosity at three diagnostic microsatellite loci that exhibited high gene-centromere meiotic recombination rates in the same family. The differences in heterozygosity levels observed with these markers were attributed to differences in the temporal application of heat shock following inert sperm activation.     

Genetic Positioning of Centromeres through Half-Tetrad Analysis in Gynogenetic Diploid Families of the Zhikong Scallop (Chlamys farreri)

Centromere mapping is a powerful tool for improving linkage maps, investigating crossover events, and understanding chiasma interference during meiosis. Ninety microsatellite markers selected across all linkage groups (LGs) from a previous Chlamys farreri genetic map were studied in three artificially induced meiogynogenetic families for centromere mapping by half-tetrad analysis. Inheritance analyses showed that all 90 microsatellite loci conformed to Mendelian inheritance in the control crosses, while 4.4 % of the microsatellite loci showed segregation departures from an expected 1:1 ratio of two homozygote classes in meiogynogenetic progeny. The second division segregation frequency (y) of the microsatellites ranged from 0.033 to 0.778 with a mean of 0.332, confirming the occurrence of partial chiasma interference in this species. Heterogeneity of y is observed in one of 42 cases in which markers were typed in more than one family, suggesting variation in gene–centromere recombination among families. Centromere location was mostly in accordance with the C. farreri karyotype, but differences in marker order between linkage and centromere maps occurred. Overall, this study makes the genetic linkage map a more complete and informative tool for genomic studies and it will also facilitate future research of the structure and function of the scallop centromeres.     

Genetic analysis of NF1: Identification of close flanking markers on chromosome 17

The gene causing von Recklinghausen neurofibromatosis, or NF1, has been more precisely localized in the pericentromeric region of chromosome 17. Narrowing of the location for the disease became possible through the identification of eight new DNA probe genetic markers in the centromeric region. Markers that closely flank the centromere also closely flank the NF1 gene. Although there was evidence against this localization in one recombinant, a review of the clinical records revealed a borderline diagnosis of NF1. Significant sex differences in recombination were observed in the pericentric region, and odds for different orders were less discriminating when sex differences were considered in multilocus analyses. The location of the NF1 gene with respect to the centromere could not be determined because recombinants between NF1 and the centromere were not detected in the set of families tested.     

Genetic mapping in hybrid zones

Recent advances in genetic mapping methodologies make it feasible to localize quantitative trait loci (QTL) that contribute to adaptation and speciation. However, it has not been possible to employ these methods in many wild species because of difficulties associated with creating and propagating recombinant populations of sufficient size for QTL mapping. Natural hybrid zones contain recombinant individuals resulting from many generations of hybridization and thus offer a potential solution to these problems. For studies of speciation, hybrid zones offer the possibility of mapping QTL simultaneously with assessments of their effects on assortative mating, hybrid fitness, and interspecific gene flow. Here, we explore the problems and prospects associated with genetic map building and QTL analyses in natural hybrid zones by analyzing correlations among markers of known genomic location in four hybrid zones between the wild sunflower species Helianthus annuus and Helianthus petiolaris. Results indicate that mapping in hybrid zones presents many challenges. These include overlap in the strength of marker correlations between linked and unlinked markers, unevenness in marker frequencies along linkages, and heterogeneity in the relationship between marker distances and correlations. All make it difficult to accurately group and order markers or to estimate the distances between them. These problems can be ameliorated by sampling strategies that maximize the difference in linkage disequilibria between linked and unlinked markers and that minimize differences in frequencies among markers or QTL. In addition, studies that employ a previously determined molecular marker map for gene localization have a greater likelihood of success than those that rely on the hybrid zone data for both map construction and QTL analyses.     

Recombination Suppression by Heterozygous Robertsonian Chromosomes in the Mouse

Robertsonian chromosomes are metacentric chromosomes formed by the joining of two telocentric chromosomes at their centromere ends. Many Robertsonian chromosomes of the mouse suppress genetic recombination near the centromere when heterozygous. We have analyzed genetic recombination and meiotic pairing in mice heterozygous for Robertsonian chromosomes and genetic markers to determine (1) the reason for this recombination suppression and (2) whether there are any consistent rules to predict which Robertsonian chromosomes will suppress recombination. Meiotic pairing was analyzed using synaptonemal complex preparations. Our data provide evidence that the underlying mechanism of recombination suppression is mechanical interference in meiotic pairing between Robertsonian chromosomes and their telocentric partners. The fact that recombination suppression is specific to individual Robertsonian chromosomes suggests that the pairing delay is caused by minor structural differences between the Robertsonian chromosomes and their telocentric homologs and that these differences arise during Robertsonian formation. Further understanding of this pairing delay is important for mouse mapping studies. In 10 mouse chromosomes (3, 4, 5, 6, 8, 9, 10, 11, 15 and 19) the distances from the centromeres to first markers may still be underestimated because they have been determined using only Robertsonian chromosomes. Our control linkage studies using C-band (heterochromatin) markers for the centromeric region provide improved estimates for the centromere-to-first-locus distance in mouse chromosomes 1, 2 and 16.