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A cellular retinoic acid-binding protein from zebrafish (Danio rerio): cDNA sequence, phylogenetic analysis, mRNA expression, and gene linkage mapping.

We report the sequence of a cDNA clone coding for a cellular retinoic acid-binding protein (CRABP) in zebrafish. The encoded polypeptide is 142 amino acids in length with an estimated molecular mass of 15.8 kDa and a calculated isoelectric point of 5.2. The zebrafish CRABP exhibits highest sequence identity to the pufferfish CRABPIIa (83%) and CRABPIIb (79%), and human CRABPII (74%) than to any other member of the intracellular lipid-binding protein (ILBP) family. A phylogenetic tree for different members of the ILBP multigene family including fatty acid-binding proteins (FABPs), cellular retinol-binding proteins (CRBPs) and CRABPs shows that the cloned zebrafish cDNA encodes a protein that clusters with CRABPs from other species and not with CRBPs and FABPs. Reverse-transcription polymerase chain reactions (RT-PCR), using oligonucleotide primers specific to the zebrafish CRABP cDNA made from total RNA of embryos collected at various developmental stages, did not detect the CRABP mRNA until 12 h post-fertilization. In adult zebrafish, CRABP mRNA was detected by RT-PCR in total RNA extracted from muscle, testes and skin, barely detectable in heart, ovary and brain and undetectable in liver, kidney and intestine. Quantitative RT-PCR (qRT-PCR) revealed a similar tissue-specific distribution for zebrafish CRABP mRNA with highest levels of CRABP mRNA in muscle followed by testes, skin, ovary and much lower levels in heart. Radiation hybrid mapping assigned the CRABP gene to linkage group 16 in the zebrafish genome. Comparison of the mapped zebrafish CRABP and human CRABPII genes revealed that zebrafish linkage group 16 has a syntenic relationship with human chromosome 1. Based on phylogenetic analysis and the syntenic relationship to the CRABPII gene in human, the zebrafish cDNA clone appears to code for a type II CRABP.

Amino Acid Sequence↗

Conserved synteny between pig chromosome 8 and human chromosome 4 but rearranged and distorted linkage maps.

The porcine genes encoding interleukin 2, alcohol dehydrogenase (class I) gamma polypeptide, and osteopontin were mapped to chromosome 8 by linkage analysis. Together with previous assignments to this chromosome (the albumin, platelet-derived growth factor receptor A, and fibrinogen genes), an extensive syntenic homology with human chromosome 4 was discovered. Loci from about three-quarters of the q arm of human chromosome 4 are on pig chromosome 8. However, the linear order of the markers is not identical in the two species, and there are several examples of interspecific differences in the recombination fractions between adjacent markers. The conserved synteny between man and the pig gives strong support to a previous suggestion that a synteny group present in the ancestor of mammalian species has been retained on human chromosome 4q. Since loci from this synteny group are found on two cattle chromosomes, the bovine rearrangement must have occurred after the split of Suidae and Bovidae within Artiodactyla.

Alcohol Dehydrogenase↗

Microsatellite genetic linkage maps of myrobalan plum and an almond-peach hybrid--location of root-knot nematode resistance genes.

Inheritance and linkage studies were carried out with microsatellite [or simple sequence repeat (SSR)] markers in a F(1) progeny including 101 individuals of a cross between Myrobalan plum ( Prunus cerasifera Ehrh) clone P.2175 and the almond (Prunus dulcis Mill.)-peach ( Prunus persica L. Batsch) hybrid clone GN22 ["Garfi" (G) almond x "Nemared" (N) peach]. This three-way interspecific Prunus progeny was produced in order to associate high root-knot nematode (RKN) resistances from Myrobalan and peach with other favorable traits for Prunus rootstocks from plum, peach and almond. The RKN resistance genes, Ma from the Myrobalan plum clone P.2175 and R(MiaNem) from the 'N' peach, are each heterozygous in the parents P.2175 and GN22, respectively. Two hundred and seventy seven Prunus SSRs were tested for their polymorphism. One genetic map was constructed for each parent according to the "double pseudo-testcross" analysis model. The Ma gene and 93 markers [two sequence characterized amplified regions (SCARs), 91 SSRs] were placed on the P.2175 Myrobalan map covering 524.8 cM. The R(MiaNem) gene, the Gr gene controlling the color of peach leaves, and 166 markers (one SCAR, 165 SSRs) were mapped to seven linkage groups instead of the expected eight in Prunus. Markers belonging to groups 6 and 8 in previous maps formed a single group in the GN22 map. A reciprocal translocation, already reported in a G x N F(2), was detected near the Gr gene. By separating markers from linkage groups 6 and 8 from the GN22 map, it was possible to compare the eight homologous linkage groups between the two maps using the 68 SSR markers heterozygous in both parents (anchor loci). All but one of these 68 anchor markers are in the same order in the Myrobalan plum map and in the almond-peach map, as expected from the high level of synteny within Prunus. The Ma and R(MiaNem)genes confirmed their previous location in the Myrobalan linkage group 7 and in the GN22 linkage group 2, respectively. Using a GN22 F(2) progeny of 78 individuals, a microsatellite map of linkage group 2 was also constructed and provided additional evidence for the telomeric position of R(MiaNem) in group 2 of the Prunus genome.

Animals↗

Linkage mapping of 40 randomly isolated liver cDNA clones in the mouse.

We report the chromosomal mapping of 43 loci for 40 randomly isolated mouse liver cDNA clones by linkage analysis in an interspecific backcross of ((C57BL/6J x Mus spretus) x C57BL/6J). The clones were sequenced from both sides and a subset was examined for expression in various mouse tissues. Fifteen of the 40 mapped cDNA clones are either identical or strongly related to known sequences in GenBank, while 25 represent new genes. Additional loci mapped in this cross include 53 simple sequence repeat polymorphisms and 40 restriction fragment length variants from previously characterized cDNA markers. Nine homologous human genes were identified for 7 mouse liver cDNA clones. One clone that maps to mouse chromosome 3 (D3Ucla1) identified a novel homologous segment (synteny) on human chromosome 18q23 (D18S372E). These studies provide linkage mapping and initial characterization of random cDNA clones that may provide a resource for the positional candidate cloning of disease genes.

Animals↗

Saturation multipoint linkage mapping of chromosome 6q in type 1 diabetes.

Linkage analysis of type 1 diabetes sib pair families (n = 334) has suggested two separate regions of human chromosome 6q are linked to disease (designated IDDM5 and IDDM8). To test if these are false positive results, all available sib pair families (n = 429) were typed using a 92% informative map of chromosome 6q and multipoint analysis. The two regions still showed positive evidence of linkage, most notably the proterminal region, 6q27, corresponding to IDDM8 (MLS = 2.57, p = 0.0006; lambda s = 1.17). In addition, some evidence of transmission disequilibrium was seen with marker a046xa9 (IDDM5).

Chromosome Mapping↗

Linkage mapping of the mouse nephrosis (nep) gene to chromosome 15.

ICGN is a partially inbred strain of mice with nephrotic syndrome caused by spontaneous glomerular lesion. It has been reported that the albuminuria in ICGN mouse was controlled by at least a single autosomal recessive gene (nep). In this study, we mapped the nep locus by linkage analysis of backcross progeny between ICGN and MSM mice using DNA pooling method. The linkage analysis revealed that the nep locus was localized on the distal part of chromosome 15.

Animals↗

A consensus linkage map for sugi (Cryptomeria japonica) from two pedigrees, based on microsatellites and expressed sequence tags.

A consensus map for sugi (Cryptomeria japonica) was constructed by integrating linkage data from two unrelated third-generation pedigrees, one derived from a full-sib cross and the other by self-pollination of F1 individuals. The progeny segregation data of the first pedigree were derived from cleaved amplified polymorphic sequences, microsatellites, restriction fragment length polymorphisms, and single nucleotide polymorphisms. The data of the second pedigree were derived from cleaved amplified polymorphic sequences, isozyme markers, morphological traits, random amplified polymorphic DNA markers, and restriction fragment length polymorphisms. Linkage analyses were done for the first pedigree with JoinMap 3.0, using its parameter set for progeny derived by cross-pollination, and for the second pedigree with the parameter set for progeny derived from selfing of F1 individuals. The 11 chromosomes of C. japonica are represented in the consensus map. A total of 438 markers were assigned to 11 large linkage groups, 1 small linkage group, and 1 nonintegrated linkage group from the second pedigree; their total length was 1372.2 cM. On average, the consensus map showed 1 marker every 3.0 cM. PCR-based codominant DNA markers such as cleaved amplified polymorphic sequences and microsatellite markers were distributed in all linkage groups and occupied about half of mapped loci. These markers are very useful for integration of different linkage maps, QTL mapping, and comparative mapping for evolutional study, especially for species with a large genome size such as conifers.

Base Sequence↗

Genomic characterization and linkage mapping of the apple allergen genes Mal d 2 (thaumatin-like protein) and Mal d 4 (profilin).

Four classes of apple allergens (Mal d 1, -2, -3 and -4) have been reported. By using PCR cloning and sequencing approaches, we obtained genomic sequences of Mal d 2 (thaumatin-like protein) and Mal d 4 (profilin) from the cvs Prima and Fiesta, the two parents of a European reference mapping population. Two copies of the Mal d 2 gene (Mal d 2.01 A and Mal d 2.01 B) were identified, which primarily differed in the length of a single intron (378 or 380 nt) and in one amino acid in the signal peptide. Both Mal d 2.01 A and Mal d 2.01 B were mapped at identical position on linkage group 9. Genomic characterization of four Mal d 4 genes (Mal d 4.01 A and B, Mal d 4.02 A and Mal d 4.03 A) revealed their complete gDNA sequences which varied among genes in length from 862 to 2,017 nt. They all contained three exons of conserved length: 123, 138, and 135 nt. Mal d 4.01 appeared to be duplicated in two copies and located on linkage group 9. Mal d 4.02 A and Mal d 4.03 A were single copy genes located on linkage group 2 and 8, respectively.

Allergens↗

A detailed linkage map of medaka, Oryzias latipes: comparative genomics and genome evolution.

We mapped 633 markers (488 AFLPs, 28 RAPDs, 34 IRSs, 75 ESTs, 4 STSs, and 4 phenotypic markers) for the Medaka Oryzias latipes, a teleost fish of the order Beloniformes. Linkage was determined using a reference typing DNA panel from 39 cell lines derived from backcross progeny. This panel provided unlimited DNA for the accumulation of mapping data. The total map length of Medaka was 1354.5 cM and 24 linkage groups were detected, corresponding to the haploid chromosome number of the organism. Thirteen to 49 markers for each linkage group were obtained. Conserved synteny between Medaka and zebrafish was observed for 2 independent linkage groups. Unlike zebrafish, however, the Medaka linkage map showed obvious restriction of recombination on the linkage group containing the male-determining region (Y) locus compared to the autosomal chromosomes.

Animals↗

Molecular genetic linkage maps for allotetraploid Leymus wildryes (Gramineae: Triticeae).

Molecular genetic maps were constructed for two full-sib populations, TTC1 and TTC2, derived from two Leymus triticoides x Leymus cinereus hybrids and one common Leymus triticoides tester. Informative DNA markers were detected using 21 EcoRI-MseI and 17 PstI-MseI AFLP primer combinations, 36 anchored SSR or STS primer pairs, and 9 anchored RFLP probes. The 164-sib TTC1 map includes 1069 AFLP markers and 38 anchor loci in 14 linkage groups spanning 2001 cM. The 170-sib TTC2 map contains 1002 AFLP markers and 36 anchor loci in 14 linkage groups spanning 2066 cM. Some 488 homologous AFLP loci and 24 anchor markers detected in both populations showed similar map order. Thus, 1583 AFLP markers and 50 anchor loci were mapped into 14 linkage groups, which evidently correspond to the 14 chromosomes of allotetraploid Leymus (2n = 4x = 28). Synteny of two or more anchor markers from each of the seven homoeologous wheat and barley chromosomes was detected for 12 of the 14 Leymus linkage groups. Moreover, two distinct sets of genome-specific STS markers were identified in these allotetraploid Leymus species. These Leymus genetic maps and populations will provide a useful system to evaluate the inheritance of functionally important traits of two divergent perennial grass species.

Amino Acid Sequence↗

High-density genetic linkage maps of Phytophthora infestans reveal trisomic progeny and chromosomal rearrangements.

Detailed analysis of the inheritance of molecular markers was performed in the oomycete plant pathogen Phytophthora infestans. Linkage analysis in the sexual progeny of two Dutch field isolates (cross 71) resulted in a high-density map containing 508 markers on 13 major and 10 minor linkage groups. The map showed strong clustering of markers, particularly of markers originating from one parent, and dissimilarity between the parental isolates on linkage group III in the vicinity of the mating-type locus, indicating a chromosomal translocation. A second genetic map, constructed by linkage analysis in sexual progeny of two Mexican isolates (cross 68), contained 363 markers and is thus less dense than the cross 71 map. For some linkage groups the two independent linkage maps could be aligned, but sometimes markers appeared to be in a different order, or not linked at all, indicating chromosomal rearrangements between genotypes. Graphical genotyping showed that some progeny contained three copies of a homologous linkage group. This trisomy was found for several linkage groups in both crosses. Together, these analyses suggest a genome with a high degree of flexibility, which may have implications for evolution of new races and resistance development to crop protection agents.

Chromosome Mapping↗

New microsatellite markers on the linkage map of the chicken genome.

Ten polymorphic microsatellite markers mainly of the poly (TG) type were examined on the two international reference families, the East Lansing reference family and the Compton reference family. Three of the markers were polymorphic on the East Lansing family (linkage groups E), three on the Compton family (linkage groups C), and four were polymorphic on both families. The microsatellite markers were well dispersed among the various linkage groups, and there was no evidence for clustering of the markers within the map. Common microsatellite markers were able to associate linkage groups from the two genetic maps. Association of linkage groups E2 with C4, E10 with C13, and E1 with C15 are found. Linkage group E1 is located on chromosome 1, and because of the association between E1 and C15, C15 is therefore assigned to chromosome 1. A newly defined linkage group (E20) was found in case of microsatellite marker MCW14. The relative distances between markers and the gene order of the markers can change upon addition of more markers to the linkage maps.

Animals↗

Physical mapping of 49 microsatellite markers on chromosome 19 and correlation with the genetic linkage map.

We have regionally localized 49 microsatellite markers developed by Généthon using a panel of previously characterized somatic cell hybrids that retain fragments from chromosome 19. The tight correlation observed between the physical and the genetic orders of the microsatellites provide cytogenetic anchorages to the genetic map data. We propose a position for the centromere just above D19S415, from the study of two hybrids, each of which retains one of the two derivatives of a balanced translocation t(1;19)(q11;q11). Microsatellites, which can be identified by a standard PCR protocol, are useful tools for the localization of disease genes and for the establishment of YAC or cosmid contigs. These markers can also judiciously be used for the characterization of new hybrid cell line panels. We report such a characterization of 11 clones, 8 of which were obtained by irradiation-fusion. Using the whole hybrid panel, we were able to define the order of 12 pairs of genetically colocalized microsatellites. As examples of gene mapping by the combined use of microsatellites and hybrid cell lines, we regionally assigned the PVS locus between the 19q13.2 markers D19S417 and D19S423 and confirmed the locations of fucosyltransferase loci FUT1, FUT2, and FUT5.

Animals↗

Development of a molecular genetic linkage map for Colletotrichum lindemuthianum and segregation analysis of two avirulence genes.

A framework genetic map was developed for the fungal pathogen Colletotrichum lindemuthianum, the causal agent of anthracnose of common bean (Phaseolus vulgaris L.). This is the first genetic map for any species within the family Melanconiaceae and the genus Colletotrichum and provides the first estimate of genome length for C. lindemuthianum. The map was generated using 106 haploid F1 progeny derived from crossing two Mexican C. lindemuthianum isolates differing in two avirulence genes (AvrclMex and AvrclTO). The map comprises 165 AFLP markers covering 1,897 cM with an average spacing of 11.49 cM. The markers are distributed over 19 major linkage groups containing between 5 and 25 markers each and the genome length was estimated to be approximately 3,241 cM. The avirulence genes AvrclMex and AvrclTO segregate in a 1:1 ratio supporting the gene for gene hypothesis for the incompatible reaction between C. lindemuthianum and P. vulgaris, but could not be incorporated into the genetic map. This initial outline map forms the basis for the development of a more detailed C. lindemuthianum linkage map, which would include other types of molecular markers and allow the location of genes previously isolated and characterized in this species.

Colletotrichum↗

An SSR- and AFLP-based genetic linkage map of tall fescue (Festuca arundinacea Schreb.).

Tall fescue (Festuca arundinacea Schreb.) is commonly grown as forage and turf grass in the temperate regions of the world. Here, we report the first genetic map of tall fescue constructed with PCR-based markers. A combination of amplified fragment length polymorphisms (AFLPs) and expressed sequence tag-simple sequence repeats (EST-SSRs) of both tall fescue and those conserved in grass species was used for map construction. Genomic SSRs developed from Festuca x Lolium hybrids were also mapped. Two parental maps were initially constructed using a two-way pseudo-testcross mapping strategy. The female (HD28-56) map included 558 loci placed in 22 linkage groups (LGs) and covered 2,013 cM of the genome. In the male (R43-64) map, 579 loci were grouped in 22 LGs with a total map length of 1,722 cM. The marker density in the two maps varied from 3.61 cM (female parent) to 2.97 (male parent) cM per marker. These differences in map length indicated a reduced level of recombination in the male parent. Markers that revealed polymorphism within both parents and showed 3:1 segregation ratios were used as bridging loci to integrate the two parental maps as a bi-parental consensus. The integrated map covers 1,841 cM on 17 LGs, with an average of 54 loci per LG, and has an average marker density of 2.0 cM per marker. Homoeologous relationships among linkage groups of six of the seven predicted homeologous groups were identified. Three small groups from the HD28-56 map and four from the R43-64 map are yet to be integrated. Homoeologues of four of those groups were detected. Except for a few gaps, markers are well distributed throughout the genome. Clustering of those markers showing significant segregation distortion (23% of total) was observed in four of the LGs of the integrated map.

Chromosome Mapping↗

Ordered subset analysis in genetic linkage mapping of complex traits.

Etiologic heterogeneity is a fundamental feature of complex disease etiology; genetic linkage analysis methods to map genes for complex traits that acknowledge the presence of genetic heterogeneity are likely to have greater power to identify subtle changes in complex biologic systems. We investigate the use of trait-related covariates to examine evidence for linkage in the presence of heterogeneity. Ordered-subset analysis (OSA) identifies subsets of families defined by the level of a trait-related covariate that provide maximal evidence for linkage, without requiring a priori specification of the subset. We propose that examining evidence for linkage in the subset directly may result in a more etiologically homogeneous sample. In turn, the reduced impact of heterogeneity will result in increased overall evidence for linkage to a specific region and a more distinct lod score peak. In addition, identification of a subset defined by a specific trait-related covariate showing increased evidence for linkage may help refine the list of candidate genes in a given region and suggest a useful sample in which to begin searching for trait-associated polymorphisms. This method provides a means to begin to bridge the gap between initial identification of linkage and identification of the disease predisposing variant(s) within a region when mapping genes for complex diseases. We illustrate this method by analyzing data on breast cancer age of onset and chromosome 17q [Hall et al., 1990, Science 250:1684-1689]. We evaluate OSA using simulation studies under a variety of genetic models.

Breast Neoplasms↗

Development of a second generation linkage map for almond using RAPD and SSR markers.

Fifty-four RAPD (random amplified polymorphic DNA) markers and 6 SSRs (simple sequence repeats) were included in a molecular marker map with 120 RFLPs (restriction fragment length polymorphisms) and 7 isozyme genes previously constructed using the offspring of a cross between the almond (Prunus amygdalus) cultivars 'Ferragnès' and 'Tuono'. Only highly reproducible RAPDs segregating 1:1 were used. To identify these markers, a total of 325 primers were screened, from which 41 produced RAPDs useful for mapping. Polymorphism was detected in six of the eight Prunus SSRs (simple sequence repeats) studied, thus enabling these to be mapped. All markers were placed on the 8 linkage groups previously identified. The number of new markers included in the map of 'Ferragnès' was 33 for a total of 126, and 30 in the map of 'Tuono' for a total of 99. The sizes of the maps of 'Ferragnès' (415 cM) and 'Tuono' (416 cM) were similar, representing a 5% increase over the maps constructed solely with isozymes and RFLPs. The estimated total size of the almond map was of 457 cM. Some markers were placed in zones with low density of markers and others in the extreme of linkage groups. The use of RAPD markers to complete genetic maps constructed with transferable markers is discussed.

Chromosome Mapping↗

Novel approaches to linkage mapping.

The recent progress seen in the development of high-quality genetic markers and high-density human genetic maps is facilitating new strategies for mapping disease-causing and disease-susceptibility genes. These strategies, which include homozygosity mapping, DNA pooling, and the utilization of linkage disequilibrium, have been successfully applied to the mapping and fine mapping of several loci causing diseases segregating in genetically isolated populations.

Chromosome Mapping↗