Detection of linkage for heterogeneous disorders by using multipoint linkage analysis.

We have compared the efficiency of the lod score test which assumes heterogeneity (lod2) to the standard lod score test which assumes homogeneity (lod1) when three-point linkage analysis is used in successive map intervals. If it is assumed that a gene located midway between two linked marker loci is responsible for a proportion of disease cases, then the lod1 test loses power relative to the lod2 test, as the proportion of linked families decreases, as the flanking markers are more closely linked, and as more map intervals are tested. Moreover, when multipoint analysis is used, linkage for a disease gene is more likely to be incorrectly excluded from a complete and dense linkage map if true genetic heterogeneity is ignored. We thus conclude that, in general, the lod2 linkage test is more efficient for detecting a true linkage when a complete genetic marker map is screened for a heterogeneous disorder.     

Tandem genetic duplications in phage lambda. IV. The locations of spontaneously arising tandem duplications.

Twenty-four spontaneously arising, long DNA addition derivatives of phage lambda have been isolated by two methods (one physical, one genetic) based on phage DNA content. All are shown to contain a tandem duplication of phage DNA by a number of criteria. The location of the duplicated segment in each has been determined by electron microscopy of DNA hereroduplexes. The duplications are found to lie at random throughout the chromosome, with no preferential locations for endpoints. This rules out the possibility that duplications are formed by crossing-over at regions of homology on the phage chromosome.     

Detection of gene-environment interactions in joint segregation and linkage analysis.

We compare approaches for analysis of gene-environment (G x E) interaction, using segregation and joint segregation and linkage analyses of a quantitative trait. Analyses of triglyceride levels in a single large pedigree demonstrate the two methods and show evidence for a significant interaction (P=.015 when segregation analysis is used; P=.006 when joint analysis is used) between a codominant major gene and body-mass index. Genotype-specific correlation coefficients, between triglyceride levels and body-mass index, estimated from the joint model are rAA=.72, rAa=.49, and raa=. 20. Several simulation studies indicate that joint segregation and linkage analysis leads to less-biased and more-efficient estimates of a G x E-interaction effect, compared with segregation analysis alone. Depending on the heterozygosity of the marker locus and its proximity to the trait locus, we found joint analysis to be as much as 70% more efficient than segregation analysis, for estimation of a G x E-interaction effect. Over a variety of parameter combinations, joint analysis also led to moderate (5%-10%) increases in power to detect the interaction. On the basis of these results, we suggest the use of combined segregation and linkage analysis for improved estimation of G x E-interaction effects when the underlying trait gene is unmeasured.     

Comparing tandem repeats with duplications and excisions of variable degree

Traditional sequence comparison by alignment employs a mutation model comprised of two events, substitutions and indels (insertions or deletions) of single positions. However, modern genetic analysis knows a variety of more complex mutation events (e.g., duplications, excisions, and rearrangements), especially regarding DNA. With ever more DNA sequence data becoming available, the need to accurately compare sequences which have clearly undergone more complicated types of mutational processes is becoming critical. Herein we introduce a new method for pairwise alignment and comparison of sequences with respect to the special evolution of tandem repeats: substitutions and indels of single positions and, additionally, duplications and excisions of variable degree (i.e., of one or more repeat copies simultaneously) are taken into account. To evaluate our method, we apply it to the spa VNTR (variable number of tandem repeats) cluster of Staphylococcus aureus, a bacterium of high medical importance     

Detection of susceptibility loci by genome-wide linkage analysis

The objective of this study is to evaluate the efficacy of a model-free linkage statistics for finding evidence of linkage using two different maps and to illustrate how the comparison of results from several populations might provide insight into the underlying genetic etiology of the disease of interest. The results obtained in terms of detection of the risk loci and threshold for declaring linkage and power are very similar for a dense SNP map and a sparser microsatellite map. The populations differed in terms of family ascertainment and diagnosis criteria, leading to different power to detect the individual underlying disease loci. Our results for the individual replicates are consistent with the disease model used in the simulation.     

Segregation and linkage analysis.

Computer programs are available in the software package SAGE to perform a variety of segregation and linkage analyses used by human geneticists. These methods are designed specifically to uncover major gene segregation in pedigree data coming from non-inbred populations. With the aid of a closely linked polymorphic marker, they can detect a locus that contributes as little as 10% to the variation of a quantitative trait in a pedigree sample of several hundred individuals.     

The effects of tandem duplications on crossing over inDrosophila melanogaster

Each of three tandem duplications,Bar, Beadex ^r49k andDp(I)z-w, when homozygous increases crossing over in their environs in excess of the genetic length of the duplication.Detailed crossing over studies withDp(I)z-w showed that in the duplication homozygotes interference is reduced and when combined with heterologous autosomal inversions, double crossovers occurring in less than 10 map units are readily recovered.These results are interpreted in terms of the concept of effective pairing and suggest that tandem duplications increase crossing over by increasing effective pairing.     

Multipoint linkage analysis.

A formula is given for the advantage of n-point sampling, which approaches infinity with n. However, 2-point and 3-point analyses extract nearly all the information in such samples and at the same time communicate this information as lods and chi 2, which can be combined with other data by simple addition without reevaluation of the likelihood. When null interference is assumed, map distances and multiple recombination frequencies are inflated, and there is substantial loss of efficiency and of support for the correct order.     

The problem of ascertainment for linkage analysis.

It is generally believed that ascertainment corrections are unnecessary in linkage analysis, provided individuals are selected for study solely on the basis of trait phenotype and not on the basis of marker genotype. The theoretical rationale for this is that standard linkage analytic methods involve conditioning likelihoods on all the trait data, which may be viewed as an application of the ascertainment assumption-free (AAF) method of Ewens and Shute. In this paper, we show that when the observed pedigree structure depends on which relatives within a pedigree happen to have been the probands (proband-dependent, or PD, sampling) conditioning on all the trait data is not a valid application of the AAF method and will result in asymptotically biased estimates of genetic parameters (except under single ascertainment). Furthermore, this result holds even if the recombination fraction R is the only parameter of interest. Since the lod score is proportional to the likelihood of the marker data conditional on all the trait data, this means that when data are obtained under PD sampling the lod score will yield asymptotically biased estimates of R, and that so-called mod scores (i.e., lod scores maximized over both R and parameters theta of the trait distribution) will yield asymptotically biased estimates of R and theta. Furthermore, the problem appears to be intractable, in the sense that it is not possible to formulate the correct likelihood conditional on observed pedigree structure. In this paper we do not investigate the numerical magnitude of the bias, which may be small in many situations. On the other hand, virtually all linkage data sets are collected under PD sampling. Thus, the existence of this bias will be the rule rather than the exception in the usual applications.     

Microsatellites for linkage analysis of genetic traits.

Microsatellites are tandem repeats of simple sequence that occur abundantly and at random throughout most eukaryotic genomes. Since they are usually less than 100 bp long and are embedded in DNA with unique sequence, they can be amplified in vitro using the polymerase chain reaction. Microsatellites are easy to clone and characterize and display considerable polymorphism due to variation in the number of repeat units. This polymorphism is sufficiently stable to use in genetic analyses. Microsatellites are therefore ideal markers for constructing high-resolution genetic maps in order to identify susceptibility loci involved in common genetic diseases.     

Sites in the AAV5 capsid tolerant to deletions and tandem duplications

Gene therapy vectors based on adeno-associated virus (AAV) have shown much promise in clinical trials for the treatment of a variety of diseases. However, the ability to manipulate and engineer the viral surface for enhanced efficiency is necessary to overcome such barriers as pre-existing immunity and transduction of non-target cells that currently limit AAV applications. Although single amino acid changes and peptide insertions at select sites have been explored previously, the tolerance of AAV to small deletions and tandem duplications of sequence has not been globally addressed. Here, we have generated a large, diverse library of >10⁵ members containing deletions and tandem duplications throughout the viral capsid of AAV5. Four unique mutants were identified that maintain the ability to form viral particles, with one showing improved transduction on both 293T and BEAS-2B cells. This approach may find potential use for the generation of novel variants with improved and altered properties or in the identification of sites that are tolerant to insertions of targeting ligands.     

Software for analysis and manipulation of genetic linkage data.

We present eight computer programs written in the C programming language that are designed to analyze genotypic data and to support existing software used to construct genetic linkage maps. Although each program has a unique purpose, they all share the common goals of affording a greater understanding of genetic linkage data and of automating tasks to make computers more effective tools for map building. The PIC/HET and FAMINFO programs automate calculation of relevant quantities such as heterozygosity, PIC, allele frequencies, and informativeness of markers and pedigrees. PREINPUT simplifies data submissions to the Centre d'Etude du Polymorphisme Humain (CEPH) data base by creating a file with genotype assignments that CEPH's INPUT program would otherwise require to be input manually. INHERIT is a program written specifically for mapping the X chromosome: by assigning a dummy allele to males, in the nonpseudoautosomal region, it eliminates falsely perceived noninheritances in the data set. The remaining four programs complement the previously published genetic linkage mapping software CRI-MAP and LINKAGE. TWOTABLE produces a more readable format for the output of CRI-MAP two-point calculations; UNMERGE is the converse to CRI-MAP's merge option; and GENLINK and LINKGEN automatically convert between the genotypic data file formats required by these packages. All eight applications read input from the same types of data files that are used by CRI-MAP and LINKAGE. Their use has simplified the management of data, has increased knowledge of the content of information in pedigrees, and has reduced the amount of time needed to construct genetic linkage maps of chromosomes.     

Methods for genetic linkage analysis using trisomies.

Certain genetic disorders are rare in the general population, but more common in individuals with specific trisomies. Examples of this include leukemia and duodenal atresia in trisomy 21. This paper presents a linkage analysis method for using trisomic individuals to map genes for such traits. It is based on a very general gene-specific dosage model that posits that the trait is caused by specific effects of different alleles at one or a few loci and that duplicate copies of "susceptibility" alleles inherited from the nondisjoining parent give increased likelihood of having the trait. Our mapping method is similar to identity-by-descent-based mapping methods using affected relative pairs and also to methods for mapping recessive traits using inbred individuals by looking for markers with greater than expected homozygosity by descent. In the trisomy case, one would take trisomic individuals and look for markers with greater than expected homozygosity in the chromosomes inherited from the nondisjoining parent. We present statistical methods for performing such a linkage analysis, including a test for linkage to a marker, a method for estimating the distance from the marker to the trait gene, a confidence interval for that distance, and methods for computing power and sample sizes. We also resolve some practical issues involved in implementing the methods, including how to use partially informative markers and how to test candidate genes.     

Continuous linkage map of human chromosome 14 short tandem repeat polymorphisms.

Nine moderately to highly informative short tandem repeat polymorphisms were assigned to chromosome 14 using somatic cell hybrids and were mapped using linkage analysis. The nine markers formed a continuous linkage map covering almost the entire long arm from 14q11.2 to q32. The markers filled a large gap within previously reported linkage maps for this chromosome. Best order of the new loci from q11.2 to q32 was D14S50, D14S54, D14S49, D14S47, D14S52, D14S53, D14S55, D14S48, and D14S51. The order shown for all adjacent pairs of loci was very strongly favored with the exception of loci pair D14S55 and D14S48, for which the order was moderately favored. Map lengths for the nine loci were 142 cM in females and 72 cM in males. Female recombination frequencies exceeded male recombination frequencies in the middle and distal portions of the map.     

Genetic instability inDrosophila melanogaster: tandem duplications as monitors of intrastrand exchange

Two tandem duplications of the X chromosome associated with the white eye color locus are described. In heterozygous females both revert to a nonduplicated chromosome without detectable meiotic recombination. Clustering of revertants suggests the reversion event occurs in the germ line prior to meiosis. Similarly in males one duplication also reverts with clustering implying a premeiotic event. Revertant X chromosomes derived from males are either nonduplicated or deleted. Intrastrand exchange can account for some but not all revertants recovered.Dedicated to Professor Bauer on the occasion of his seventy-fifth birthday