Mapping of rat chromosome 5 markers generated from chromosome-sorted DNA

Nineteen markers for rat Chromosome 5 (Chr) were generated by screening chromosome-sorted DNA libraries and were subsequently mapped by linkage to known markers by use of five F₂ rat populations. Along with existing markers, these newly produced markers are potentially useful for fine mapping of certain quantitative trait loci for blood pressure and for obesity. Received: 20 January 1997 / Accepted: 17 March 1997     

Linkage mapping of rat chromosome markers generated from chromosome-sorted DNA

Ninety-one new rat microsatellite chromosome markers were generated through screening chromosome-sorted DNA libraries. Of the 91 markers, 29 have been mapped to various rat chromosomes. Because of a lack of suitable polymorphisms among the appropriate rat strains of our interest, the remaining 62 markers are still unassigned, but are likely to be useful for genotyping different rat strains employed to study a wide range of genetic traits other than blood pressure. With these new markers, two genes, encoding α₂ adrenergic receptor, class II and gastric H,K-ATPase beta subunit, were mapped to regions on rat Chromosomes (Chrs) 1 and 16 respectively. Received: 2 July 1997 / Accepted: 13 September 1997     

Linkage mapping of rat Chromosome 5 markers generated from chromosome-specific libraries

Seventy-six novel microsatellite markers with various simple sequence repeat (SSR) motifs are reported in this paper. They were generated on the basis of non-radioactive library screening procedures from flow-sorted rat Chromosome (Chr) 5-specific DNA, and were mapped in three rat backcross populations. Fifty-four of these markers mapped to Chr 5, while the other 22 mapped to other chromosomes of the rat genome. The marker D3Uwm8 is a new microsatellite marker for the rat syndecan 4 (ryudocan) gene. A genotyping protocol based on agarose gel electrophoresis is also provided in this paper. Received: 17 December 1998 / Accepted: 17 February 1999     

Mapping of rabbit chromosome 1 markers generated from a microsatellite-enriched chromosome-specific library

A genomic DNA library was produced from flow-sorted rabbit chromosome 1 and enriched for fragments containing CA-repeats. Clones containing CA-repeats were identified and primers for amplification of the microsatellite were developed after sequencing the clone. The degree of polymorphism was tested in rabbits from different breeds. This approach identified 12 microsatellite markers which could be used for studying linkage relationships in the progeny of an F(2)-intercross: (AX/JUxIIIVO/JU) F(2), and two backcrosses: (OS/JxX/J)X/J and (WH/JxX/J)X/J. Seven of these markers were mapped on chromosome 1.     

Statistical analysis of binary data generated from multilocus dominant DNA markers

The use of methodologies such as RAPD and AFLP for studying genetic variation in natural populations is widespread in the ecology community. Because data generated using these methods exhibit dominance, their statistical treatment is less straightforward. Several estimators have been proposed for estimating population genetic parameters, assuming simple random sampling and the Hardy-Weinberg (HW) law. The merits of these estimators remain unclear because no comparative studies of their theoretical properties have been carried out. Furthermore, ascertainment bias has not been explicitly modelled. Here, we present a comparison of a set of candidate estimators of null allele frequency (q), locus-specific heterozygosity (h) and average heterozygosity () in terms of their bias, standard error, and root mean square error (RMSE). For estimating q and h, we show that none of the estimators considered has the least RMSE over the parameter space. Our proposed zero-correction procedure, however, generally leads to estimators with improved RMSE. Assuming a beta model for the distribution of null homozygote proportions, we show how correction for ascertainment bias can be carried out using a linear transform of the sample average of h and the truncated beta-binomial likelihood. Simulation results indicate that the maximum likelihood and empirical Bayes estimator of have negligible bias and similar RMSE. Ascertainment bias in estimators of is most pronounced when the beta distribution is J-shaped and negligible when the latter is inverse J-shaped. The validity of the current findings depends importantly on the HW assumption-a point that we illustrate using data from two published studies.     

A single linkage group comprising 11 polymorphic DNA markers on rat Chromosome 3

Eleven polymorphic DNA markers were mapped to rat Chromosome (Chr) 3 by linkage analysis of F₂ progeny of F344/N and LEW/N rat strains. The markers, including seven genes and four anonymous loci, formed a single linkage group covering approximately 112 cM with the following order: Ptgs1 (prostaglandin G/H synthase I)-D3Arb178-Scn2a (sodium channel, type II, -polypeptide)-D3Arb1-Cat (catalase)-Bdnf (brain-derived neurotrophic factor)-D3Arb219-D3Arb2-Sus2 (seminal vesicle secretion II protein)-Sdc4 (ryudocan/syndecan4)-Stnl (statin-like protein). Eight of these markers were analyzed for polymorphisms in 14 additional inbred rat strains. Three to five alleles were detected for each marker, suggesting that they are highly polymorphic and useful for genetic mapping studies with inbred rat strains. Chromosomal syntenic conservation among rats, mice and humans is also discussed.     

Mapping of 55 new rat microsatellite markers from chromosome-specific libraries

Fifty-five novel rat microsatellite markers were isolated from libraries specific for rat chromosomes (Chrs) 1, 2, and 7. The markers were mapped in three backcross rat populations. Thirty of these markers mapped to Chrs 1, 2, or 7, while the other 25 mapped to other chromosomes. New markers for two genes, liver-specific transporter gene (Livtr) and insulin-responsive glucose transporter (Glut4), were also mapped to rat Chrs 9 and 10, respectively. Three provisionally assigned markers from previous studies were also confirmed. Detailed methodologies for the generation and enrichment of clones containing repeat sequences and for the isolation of chromosome-specific markers are presented, since they represent unique combinations and modifications of previous protocols. Such methods and the newly presented markers should be useful for both specific and general mapping studies in the rat. Received: 11 February 1998 / Accepted: 7 April 1998     

Mapping of porcine genetic markers generated by representational difference analysis

Representational difference analysis (RDA) was performed using pig genomic DNA from a Landrace non-selected control population and a Landrace population selected for increased loin muscle area (LMA) for five generations. Pigs used for the analysis differed phenotypically for various carcass traits and were divergent in genotype at the skeletal muscle ryanodine receptor 1 locus. Two RDA experiments were performed using BamHI and BglII. Fourteen BamHI and 37 BglII difference products were cloned and sequenced. Oligonucleotide primers were designed to amplify RDA difference products and sequence-tagged sites (STS) were developed for 16 RDA fragments (two BamHI and 14 BglII). These 16 STS were mapped using the INRA-Minnesota porcine Radiation Hybrid panel. Polymorphisms identified in nine of the STS were used to place these markers on the PiGMaP genetic linkage map. Sequence-tagged sites were localized to 11 different chromosomes including three markers on chromosome 11 and four markers on chromosome 14. Development of RDA markers increases the resolution of the pig genome maps and markers located within putative quantitative trait locus (QTL) regions can be used to refine QTL positions.     

A genetic map of microsatellite markers on rat Chromosome 7

Nine microsatellite loci were mapped to rat Chromosome (Chr) 7 by genetic linkage and somatic cell hybrid analysis. These loci include the gene encoding a member of the IID sub-family of cytochrome P450 (Cyp2d), a gene with repetitive sequences expressed during myotube formation (D7Arb1e), four anonymous loci, D7Arb81, D7Arb208, D7Arb569, D7Arb609a, and three DNA loci defined by MapPair^TM markers R245, R513, and R1071. The nine loci were all identified by PCR-based microsatellite polymorphism analysis and were characterized in 40 F₂ intercross progeny of Fischer (F344/N) and Lewis (LEW/N) rats for segregation analysis. These markers formed a single linkage group spanning 76.8 cM with the following order and distances: D7Arb569-11.4 cM-D7Arb81-9.7 cM-R513-2.6 cM-Cyp2d-0.0 cM-R245-1.3 cM-D7Arb1e-10.4 cM-R1071-15.9 cM-D7Arb609a-15.4 cM-D7Arb208. Physical mapping of Cyp2d by somatic cell hybrid analysis allowed us to assign this linkage group to rat Chr 7. For each marker, two to six alleles were detected in a panel of 16 inbred rat strains (ACI/N, BN/SsN, BUF/N, DA/Bkl, F344/N, LER/N, LEW/N, LOU/MN, MNR/N, MR/N, SHR/N, SR/Jr, SS/Jr, WBB1/N, WBB2/N, WKY/N).     

Linkage map of seven polymorphic markers on rat Chromosome 18

A genetic linkage map of seven polymorphic markers was created with F₂ intercross progeny of F344/N and LEW/N rats and assigned to rat Chromosome (Chr) 18. Five of the markers described were defined by simple sequence length polymorphisms (SSLPs) associated with five genes: transthyretin (TTR), trypsin inhibitor-like protein (TILP), 2 adrenergic receptor (ADRB2), olfactory neuron-specific G protein (OLF), and gap junction protein (GJA1). One marker was defined by a restriction fragment length polymorphism (RFLP) detected with a probe for the human colony stimulating factor 1 receptor (CSF1R) gene. The D18N1R locus was defined by an anonymous DNA fragment amplified by the randomly amplified polymorphic DNA (RAPD) technique with a single short primer. These seven DNA loci formed a single genetic linkage group 30.4 cM in length with the following order: TTR-6.8 cM-D18N1R-9.1 cM-TILP-4.3 cM-CSF1R-0 cM-ADRB2-10.2 cM-OLF-0 cM-GJA1. The five SSLP markers were highly polymorphic. In a total of 13 inbred rat strains analyzed (F344/ N, LEW/N, LOU/MN, WBB1/N, WBB2/N, MR/N, MNR/N, ACI/N, SHR/N, WKY/N, BN/SsN, BUF/N, and LER/N), three to six alleles were detected for each marker. Remarkable linkage conservation was detected between the region of rat Chr 18 mapped and a region of mouse Chr 18. However, genes associated with these markers have been mapped to three different human chromosomes (Chrs 5, 6, and 18). The markers described here should be useful for genetic mapping studies and genetic monitoring of inbred rat strains.     

Mapping of a novel MEN-like syndrome locus to rat Chromosome 4

Multiple endocrine neoplasia-like syndrome (MENX) is a hereditary cancer syndrome in the rat characterized by inborn cataract and multiple tumors affecting the neuroendocrine system developed within the first year of life. The spectrum of affected organs is intermediate between MEN type 1 (MEN1) and MEN type 2 (MEN2) syndromes in human, but, in contrast to them, MENX is inherited in a recessive fashion. Here we report the mapping of the MENX locus to rat Chromosome (Chr) 4 by a genome-wide linkage analysis. This analysis was done in 41 animals obtained from a (Wistar/Nhg × SD^we) × SD^we interstrain backcross, where SD^we (Sprague-Dawley white eye) indicates the affected animals. The MENX disease locus was ultimately mapped to a ~22-cM interval on Chr 4 that includes the rat homolog of the human RET proto-oncogene. As activating point mutations of RET are known to be responsible for MEN2 in human, we analyzed several markers located in the proximity of Ret for linkage to the disease phenotype. Our data exclude Ret involvement in MENX and establish that a second gene, playing a role in endocrine tumor formation, lies within the distal part of rat Chr 4. Although heritable human endocrine tumors are quite rare, sporadic tumors of MEN-affected tissues occur at a much higher frequency, and their pathogenesis is poorly understood. The identification of the MENX gene should contribute to our understanding of the genetic mechanisms of neuroendocrine tissue tumorigenesis and may assist in developing new and more appropriate therapeutic strategies for these diseases.     

Map of seven polymorphic markers on rat Chromosome 14: linkage conservation with human Chromosome 4

Seven polymorphic markers identified by polymerase chain reaction (PCR) amplification, including markers for six genes—DRD1L (dopamine receptor, D1-like-2), GLUKA (glucokinase), PF4 (platelet factor 4), ALB (albumin), AFP (-fetoprotein), and BSP (bone sialoprotein)—and one anonymous locus (D14N52), were mapped to a single 67-cM linkage group with F₂ intercross progeny of F344/N and LEW/N inbred rat strains. Two of these markers, ALB and AFP, have previously been assigned to rat Chromosome (Chr) 14, allowing assignment of this entire linkage group. Five of the markers—DRD1L, PF4, ALB, AFP, and PBSP—have been physically mapped to a large region of human Chr 4 encompassing the p arm and the q arm to band q28. Homologs of two of the markers, ALB and AFP, have been mapped to Chr 5 in the mouse. Comparison of human Chr 4 with the homologous regions on Chr 14 of the rat and Chr 5 of the mouse indicated that linkage conservation with human Chr 4 extends over a greater region in the rat than in the mouse. The markers described here were found to be highly polymorphic in twelve inbred strains (F344/N, LEW/N, ACI/N, BUF/N, BN/SsN, LOU/MN, MNR/N, MR/N, SHR/N, WBB1/N, WBB2/N, and WKY/N). These polymorphic markers should be useful in genetic linkage studies of important phenotypes in rats.     

Mapping dominant markers using F2 matings

The development of efficient methods for amplifying random DNA sequences by the polymerase chain reaction has created the basis for mapping virtually unlimited numbers of mixed-phase dominant DNA markers in one population. Although dominant markers can be efficiently mapped using many different kinds of matings, recombination frequencies and locus orders are often mis-estimated from repulsion F₂ matings. The major problem with these matings, apart from excessive sampling errors of recombination frequency () estimates, is the bias of the maximum-likelihood estimator (MLE) of ( _ML). when the observed frequency of double-recessive phenotypes is 0 and the observed frequency of double-dominant phenotypes is less than 2/3 — the bias for those samples is — . We used simulation to estimate the mean bias of _ML. Mean bias is a function of n and and decreases as n increases. Valid maps of dominant markers can be built by using sub-sets of markers linked in coupling, thereby creating male and feamle coupling maps, as long as the maps are fairly dense (about 5 cM) — the sampling errors of increase as increases for coupling linkages and are equal to those for backcross matings when =0. The use of F₂ matings for mapping dominant markers is not necessarily proscribed because they yield twice as many useful markers as a backcross population, albeit in two maps, for the same number of DNA extractions and PCR assays; however, dominant markers can be more effeciently exploited by using doubled-haploid, recombinant-inbred, or other inbred populations.