A note on the combination of estimates of a recombination fraction

A number of ways of combining two or more independent estimates of the same recombination fraction can be found in the literature. We revisit this topic in the context of human gene mapping, and explore the value of transforming the recombination fraction to a new parameter whose log-likelihood function is more nearly quadratic. It is shown that the arcsine of the cube-root is one such function. These observations lead naturally to a way of summarizing and combining the summarized set of log-likelihood functions of a common recombination fraction. This idea is illustrated using pedigree data concerning six loci on chromosome 10 from the CEPH consortium. A comparison is also made with the method of summarizing and combining using 'equivalent numbers' of recombinants and informative meioses.     

Gene localization in a family with X-linked syndromal mental retardation (Prieto syndrome).

A mapping study was performed on a 3-generation Spanish family with X-linked syndromal mental retardation. Affected males have a typical facial appearance, ear malformations, abnormal growth of teeth, clinodactyly, dimpled skin at the lower back, and patellar luxation. In pneumoencephalography a marked subcortical cerebral atrophy was evident. In the linkage studies with polymorphic DNA markers, no recombination was found between the disease locus and the loci OTC and DXS148, both assigned to Xp21.1. One or more recombinants were observed between the disease locus and loci from the distal part of Xp and the pericentromeric region. Close linkage to loci of Xq has also been excluded. The analysis of multiple informative meioses suggests that the disease locus maps between DXS255 (Xp11.22) and DXS84 (Xp21.1) on Xp.     

Large-scale integration of human genetic and physical maps

Genetic maps are used routinely in family-based linkage studies to identify the rough location of genes that influence human traits and diseases. Unlike physical maps, genetic maps are based on the amount of recombination occurring between adjacent loci rather than the actual number of bases separating them. Genetic maps are constructed by statistically characterizing the number of crossovers observed in parental meioses leading to the transmission of alleles to their offspring. Considerations such as the number of meioses observed, the heterozygosity and physical distance between the loci studied, and the statistical methods used can impact the construction and reliability of a genetic map. As is well known, poorly constructed genetic maps can have adverse effects on linkage mapping studies. With the availability of sequence-based maps, as well as genetic maps generated by different researchers (such as those generated by the Marshfield and deCODE groups), one can investigate the compatibility and properties of different maps. We have integrated information from the most current human genome sequence data (UCSC genome assembly Human July 2003) as well as 8399 microsatellite markers used in the Marshfield and deCODE maps to reconcile the these maps. Our efforts resulted in updated sex-specific genetic maps.     

Genetic linkage analysis identifies new proximal and distal flanking markers for the X-linked agammaglobulinemia gene locus, refining its localization in Xq22

Genetic linkage analysis has been instrumental in mapping thegene for X-linked agammaglobulinemia (XLA) to the proximal longarm of the human X chromosome, to Xq22. Due to the relativerarity of this disease the localization of the gene within Xq22has remained imprecise. We have investigated twenty-nine familiesaffected by XLA and have found no recombinants with the DXS178locus in over 30 informative meioses. DXS178 is now the mostreliable and informative locus for use in pre-natal diagnosisand carrier detection of XLA. In addition, we have identifiednew closely linked proximal and distal flanking markers forXLA, DXS442 and DXS101, respectively. These loci are separatedby 2cM, considerably reducing the extent of DNA within whichthe XLA locus can be contained. This will open up the way formore directed positional cloning efforts for the isolation ofthe XLA gene.     

Additions to the myotonic dystrophy linkage group

One hundred and thirty members of a family with 33 cases of Dystrophia myotonica (Dm) were tested for various genetic markers including Km, Jk, Lu and Se. All individuals tested were Lua negative, but many were heterozygous for the other markers. Except for two cases all patients with Dm were positive for Km3 and Jka and they were secretors. Assuming the genes are linked and that the order of the loci is Km, Jk, Lu, Se, Dm, then there were 22 meioses informative for Dm and in 10 of these a recombination must have occurred between two of the five loci. Looking at individual pairs of loci, there were no recombinations between Km and Jk in seven informative meioses, three recombinations out of 10 meioses informative for Jk/Se, no recombinations between Se and Dm in five informative meioses and three recombinations out of 12 meioses informative for Jk/Dm.     

New recombinants within the MHC (B-complex) of the chicken

In a search for genetic recombinations within the major histocompatibility complex (MHC) of the chicken, the B-complex, the offspring from matings between heterozygous B15/B21 and B4/B6 animals were analysed by red cell agglutination. Among the progeny, 8,912 informative typings were performed. Four recombinants were found, all separating the B-complex loci B-F and B-G (B-F codes for Class I antigens, B-G codes for an antigen of which there is no known homologue in mammals). B-L (Class II antigen) always followed B-F. Stimulation in graft versus host reactions and in mixed lymphocyte cultures followed B-F/B-L. The mapping distance between the two loci B-F and B-G is in the range of 0.04 centimorgan. The lack of recombinants separating individual B-F loci in this study and in the studies of others might indicate that chicken MHC is less complex than those of mammalian species, but alternative explanations are also possible. So far no serologically defined recombinant separating Class I (B-F) and Class II (B-L) loci has been found.     

The locus ordering problem

Studies of phenotypes defined by codominant alleles at two or more loci in three-generation families allow haplotypes to be deduced. These data are easily summarized by the Mendelian convention of upper and lower case, with case defining phase rather than nature. In two-generation families haplotypes may be inferred with high precision for closely linked loci even if an allele at one locus is recessive.
Coding procedures are discussed and a simple solution to inferring the most likely order of three or more loci, and defining its likelihood compared with other orders, is presented.     

The population genetics of Duchenne: natural and artificial selection in Duchenne muscular dystrophy.

The dynamics of X linkage was derived by Haldane in 1935. It is clear that the majority of mutations to an X linked lethal are new and that methods of control based on relatives of known cases can have limited impact on future incidence. The ability to define and track neighbouring loci allows some carriers who are daughters of carriers to be detected, and possible carriers to be excluded, with high reliability. Fetal diagnosis may also be made in the same way, but not without a substantial casualty rate. The precision of such diagnosis by proxy is limited both by the estimate of the recombination fraction and its variance, and can rarely exceed 1/s where the recombinational data are based on s informative meioses. Bracketing loci provide greater security from failure to diagnose cases but may involve substantial casualty rates. The estimation of both failure rates and casualty rates is discussed.     

Reliability theory and clinical psychology

Psychologists who report test results usually ignore the published data on the reliability of the test. When reliability is taken into account, the standard error of measurement is used to estimate the magnitude of the range around the observed score in which the true score is most likely to occur. It is assumed that the observed score is the best available estimate of the true score and that the standard error of measurement is the standard deviation of possible true scores around this observed score that could result from errors of measurement. Most textbooks of psychometrics notwithstanding, neither of these popular beliefs is correct. The correct procedure for estimating the most probable true score and the range of error associated with it is described, and the implications for psychological report writing are discussed.     

Assessment of risks in connection with use of a recombinant E. coli strain for production of human growth hormone

A number of studies were performed with the aim of elucidating possible risks in connection with the use of a recombinant E. coli strain for the production of human growth hormone (hGH). The survival of recombinant E. coli in the gastrointestinal tract in rats was studied. The results showed that the recombinants were only detectable in the faeces one to two days after the administration of 10(11) bacteria. Since the recombinants are resistant to ampicillin, survival in the gastrointestinal tract was improved in rats treated with ampicillin. However, no recombinants could be detected a few days after the discontinuation of ampicillin treatment. Studies of transfer of plasmid DNA from recombinant E. coli to other bacteria strongly indicated that the possibility of transfer of the recombinant plasmid pHD117 used in the production of hGH to enterobacteria in rats is minimal. In vitro experiments have shown that transfer by conjugation, presumably after recombination between a non mobilizable plasmid such as pHD117 and a conjugative plasmid, can be calculated to be 4 X 10(-6). An uptake of free plasmid DNA in bacteria was only observed when the bacteria were treated with CaCl2. The evaluation of risks in connection with inadvertent consumption of the hGH producing bacteria has included studies of the absorption of hGH after peroral administration to rats. Approximately 0.005% was found to be absorbed. A prerequisite for absorption of hGH after consumption of recombinants is a lysis of the bacteria.(ABSTRACT TRUNCATED AT 250 WORDS)     

Rapid efficient production of baculovirus expression vectors.

Recombinant baculoviruses have been used to produce foreign proteins and have the potential to be safe, efficacious insecticides. Isolation of recombinant virus is usually by plaque phenotype. Typical recombination rates are less than 1%, thus requiring time consuming inspection of hundreds of individual plaques. We describe a method of generating recombinants which requires less time than current protocols and frequently produces recombinants at rates exceeding 30%. This protocol employs liposome-mediated transfection, reduced post-transfection incubation times, linearized parental virus which produces occlusion positive plaques in clones of the parental genotype, and colorimetric detection of recombinants. This protocol allows the initial, and frequently the final, isolation of recombinants in 7 days.     

Intertypic recombination in poliovirus: genetic and biochemical studies

Poliovirus strains of type 1 and type 3 carrying genetically mapped ts mutations and differring in growth response to guanidine have been used to infect HeLa cells. With four heterotypic pairs of the mutants, recombinants with the crossover points between the loci coding for the antigenic properties, on the one hand, and for the sensitivity to guanidine, on the other, have been obtained. The recombinants have been identified on the basis of their phenotypic properties and, in particular, of the pattern of inheritance of unselected markers. One recombinant has been characterized by fingerprinting virus-specific polypeptides. It has been found that the capsid proteins (VP2, VP3, and VP1) of this recombinant originate from the type 3 parent, whereas the nonstructural polypeptides (X, 2, and 4) are inherited from the type 1 parent. Implications of the poliovirus intertypic recombination are discussed.     

Characterization of intertypic recombinants of the Epstein-Barr virus from the body-cavity-based lymphomas cell lines BC-1 and BC-2

Epstein-Barr Virus (EBV) can infect and transform human B-lymphocytes and has been associated with numerous human malignancies. Two distinct types of EBV have been described, EBV-1 and EBV-2. Whereas type 1 is known to be most widespread throughout the healthy adult population, type 2 EBV has been shown to be significantly present in certain T-cell immunocompromised patients. Some evidence also suggests that such immune impairment promotes coinfection with multiple strains of EBV and fosters the development of intertypic recombinant viruses. In this work, we have analyzed two established body-cavity-based lymphoma or primary effusion lymphoma cell lines, BC-1 and BC-2, for the presence of intertypic EBV recombinants. Using PCR primers to amplify across several markers in the genome, we have typed the BC-1 and BC-2 EBV at these loci. Immunoblot analysis of the EBNA1 protein expressed by these cell lines also suggests a change in EBV typing at this locus in these genomes. Additionally, we have analyzed the expression patterns of the latent EBNA proteins from these viruses and performed Southern blot analysis of the BamHI- and EcoRI-digested genomes to detect variations occurring from type I and II genomes. On the basis of these data, we suggest that the genomes of EBV in BC-1 and BC-2 are intertypic recombinants of type 1 and type 2 EBV genomes. This work corroborates other reports that intertypic EBV recombinants occur in the immunocompromised population. It is likely that intertypic recombination is a mechanism by which novel variants of EBV emerge having selective advantages over a strictly type 1 or type 2 strain.     

Studies of biological properties of the recombinants between human influenza and fowl plague viruses as related to genome composition

Some biological properties of recombinants obtained by crossing of fowl plague and human influenza viruses were studied. The capacity of the recombinants to reproduce in chick embryo fibroblast cultures was in reverse correlation to the number of genes coding for P proteins derived from the human influenza virus. The genome composition was of importance for the expression of ts-phenotype of the recombinants in different systems. Substitution of at least one gene in the fowl plague virus genome by a corresponding human influenza virus gene resulted in the decrease of virulence for 1-day-old chickens. The presence of three P genes from human influenza virus genome in the genome of the recombinant proved to be insufficient for the capability of the recombinant to reproduce in organ cultures of human origin.     

New X-linked mental retardation syndrome with the gene mapped tentatively in Xp22.3

X-linked mental retardation (XLMR) is genetically heterogeneous and clinically variable. We describe a new XLMR syndrome of severe mental retardation and multiple congenital anomalies. Two sisters have (with 3 different partners) 3 severely handicapped sons. In 2 cases, oligohydramnios and intrauterine growth retardation were noted. Common anomalies included a square-shaped face, high and broad forehead, frontal bossing, downward slant of palpebral fissures, hypertelorism, epicanthic folds, long philtrum, thin upper lip, and apparently low-set ears. One boy has bilateral microphthalmos and sclerocornea, and his cousin has atrophy of the optic nerve. All 3 patients are blind and have profound statomotor and mental retardation, seizures, and a grossly abnormal electroencephalographic pattern. Additional findings are short stature, delayed bone maturation, hydronephrosis, vesicorenal reflux, cryptorchidism, clinodactyly of the 5th fingers, and transverse palmar creases. The karyotype is normal (46,XY). Segregation analysis showed perfect coinheritance between the clinical phenotype and alleles at several loci in Xp22.3, whereas recombinants were identified with marker loci from Xp22.2-qter. Analysis of multiple informative meioses suggests that the disease locus maps in Xp22.3 distal to DXS16. © 1996 Wiley-Liss, Inc.     

Frequent isolation of intertypic poliovirus recombinants with serotype 2 specificity from vaccine-associated polio cases.

Five representatives from a collection of 21 Sabin type 2-like poliovirus strains isolated from paralytic poliomyelitis cases in two regions of the USSR have been subjected to limited nucleotide sequencing. All proved to be intertypic recombinants having the genes encoding capsid proteins of Sabin 2 origin and a 3'-end portion of the genome derived from either type 1 (3 isolates) or type 3 (2 isolates) Sabin strains. The crossover points in all the 5 genomes have been mapped to different loci of the P3 region. At least 6 additional isolates from the same collection (and 2 isolates from healthy contacts), appeared to have a type 2/type 1 recombinant genome, as judged by oligonucleotide mapping. The biological significance of frequent occurrence of recombinants among field isolates of vaccine-related strains is discussed.     

Studies on the recombination between RNA genomes of poliovirus: the primary structure and nonrandom distribution of crossover regions in the genomes of intertypic poliovirus recombinants

A series of intertypic (type 3/type 1) poliovirus recombinants was obtained whose crossover sites were expected to be located in the middle of the viral genome, between the loci encoding type-specific antigenic properties, on the 5' side, and an altered sensitivity to guanidine, on the 3' side. The primary structures of the crossover regions in the genomes of these recombinants were determined by the primer extension method. The length of the crossover sites (the uninterrupted sequences shared by the recombinant and both parental genomes that are flanked, in the recombinant RNAs, by two heterotypic segments) varied between 2 and 32 nucleotides, but the majority of the sites were 5 nucleotides long or shorter. The crossover sites were nonrandomly distributed over the presumably available genome region: only a single such site was found within the gene for polypeptide 2A, whereas an apparent clustering of the crossover sites was encountered in other genomic segments. When the crossover sites were superimposed on a model of the secondary structure of the relevant region of the viral RNA molecule, a pattern consistent with the previously proposed mechanism of poliovirus recombination (L.I. Romanova, V.M. Blinov, E.A. Tolskaya, E.G. Viktorova, M.S. Kolesnikova, E.I. Guseva, and V.I. Agol (1986) Virology 155, 202-213) was observed. It is suggested that the nonrandom distribution of the crossover sites in the genomes of intertypic poliovirus recombinants was due to two factors: the existence of preferred sites for recombination, and selection against recombinants with a lowered level of viability.     

The LW: C3 recombination fraction in female meioses

No recombinants between LW and C3 , using a C3 DNA probe, were observed in 16 female meioses: z 4·216 at θ= 0·00. Combined with the data of Sistonen (1984) the recombination fraction between LW and C3 is estimated to be 0.09 ( 3·773) in females.     

High-resolution HLA haplotype frequencies of stem cell donors in Germany with foreign parentage: How can they be used to improve unrelated donor searches?

In hematopoietic stem cell transplantation, human leukocyte antigens (HLA), usually HLA loci A, B, C, DRB1 and DQB1, are required to check histocompatibility between a potential donor and the recipient suffering from a malignant or non-malignant blood disease. As databases of potential unrelated donors are very heterogeneous with respect to typing resolution and number of typed loci, donor registries make use of haplotype frequency-based algorithms to provide matching probabilities for each potentially matching recipient/donor pair. However, it is well known that HLA allele and haplotype frequencies differ significantly between populations. We estimated high-resolution HLA-A, -B, -C, -DRB1 haplotype and allele frequencies of donors within DKMS German Bone Marrow Donor Center with parentage from 17 different countries: Turkey, Poland, Italy, Russian Federation, Croatia, Greece, Austria, Kazakhstan, France, The Netherlands, Republic of China, Romania, Portugal, USA, Spain, United Kingdom and Bosnia and Herzegovina. 5-locus haplotypes including HLA-DQB1 are presented for Turkey, Poland, Italy and Russian Federation. We calculated linkage disequilibria for each sample. Genetic distances between included countries could be shown to reflect geography. We further demonstrate how genetic differences between populations are reflected in matching probabilities of recipient/donor pairs and how they influence the search for unrelated donors as well as strategic donor center typings.     

Differentiation of Potato virus Y strains using improved sets of diagnostic PCR-primers

Potato virus Y (PVY) is one of the most important viruses of potato world-wide, several strain groups are recognized. In the past two decades, novel PVY variants have appeared causing necrotic symptoms on potato tubers. Implicated are two groups of recombinant strains: PVY^NW and PVY^NTN, and NA-PVY^NTN. While the first two are recombinants between PVY-N- and O-strains the latter is a recombinant between an N-strain and an unknown PVY strain or other Potyvirus. Available biological and molecular data on PVY suggest that classification of PVY strains has to be revised. Some drawbacks have been found with recently published primers used in RT-PCR based differentiation of PVY strains as some defined isolates could not be identified correctly. Consequently we developed new primers using both recently available sequences and newly generated complete sequences of PVY strains. The reliability of these newly developed primers and procedures was successfully demonstrated on nearly 100 biologically and serologically characterised PVY isolates.