A genetic map of Peromyscus with chromosomal assignment of linkage groups (a Peromyscus genetic map).

The rodent genus Peromyscus is the most numerous and species-rich mammalian group in North America. The naturally occurring diversity within this genus allows opportunities to investigate the genetic basis of adaptation, monogamy, behavioral and physiological phenotypes, growth control, genomic imprinting, and disease processes. Increased genomic resources including a high quality genetic map are needed to capitalize on these opportunities. We produced interspecific hybrids between the prairie deer mouse (P. maniculatus bairdii) and the oldfield mouse (P. polionotus) and scored meiotic recombination events in backcross progeny. A genetic map was constructed by genotyping of backcross progeny at 185 gene-based and 155 microsatellite markers representing all autosomes and the X-chromosome. Comparison of the constructed genetic map with the molecular maps of Mus and Rattus and consideration of previous results from interspecific reciprocal whole chromosome painting allowed most linkage groups to be unambiguously assigned to specific Peromyscus chromosomes. Based on genomic comparisons, this Peromyscus genetic map covers ~83% of the Rattus genome and 79% of the Mus genome. This map supports previous results that the Peromyscus genome is more similar to Rattus than Mus. For example, coverage of the 20 Rattus autosomes and the X-chromosome is accomplished with only 28 segments of the Peromyscus map, but coverage of the 19 Mus autosomes and the X-chromosome requires 40 chromosomal segments of the Peromyscus map. Furthermore, a single Peromyscus linkage group corresponds to about 91% of the rat and only 76% of the mouse X-chromosomes.

A unique late-replicating XY to autosome translocation in Peromyscus melanophrys.

We report on the characterization of the Peromyscus melanophrys karyotype and sex chromosome system. Classic studies reported the sex chromosome system of this species may be as complex as an X(1)X(1)X(2)X(2)/X(1)X(2)Y(1)Y(2) and provided conflicting identification of the X chromosome. Using Peromyscus maniculatus chromosome paints, we have positively identified the sex chromosomes and clarified the sex determining system that once perplexed Peromyscus researchers. The sex chromosomes are characterized by a unique autosomal translocation of DNA shared between both the X and Y chromosomes. The translocated material is late replicating and heterochromatic yet retains the active chromatin conformation. Thus, autosomal regions derived from translocations involving repeat-rich material may retain some epigenetic marks specific to the sex chromosomes despite loss of epigenetic silencing activity.

Five Hundred Microsatellite Loci for Peromyscus.

Mice of the genus Peromyscus, including several endangered subspecies, occur throughout North America and have been important models for conservation research. We describe 526 primer pairs that amplify microsatellite DNA loci for P. maniculatus bairdii, 467 of which also amplify in P. polionotus subgriseus. For 12 of these loci, we report diversity data from a natural population. These markers will be an important resource for future genomic studies of Peromyscus evolution and mammalian conservation.

Comparative genome mapping of the deer mouse (Peromyscus maniculatus) reveals greater similarity to rat (Rattus norvegicus) than to the lab mouse (Mus musculus).

BACKGROUND: Deer mice (Peromyscus maniculatus) and congeneric species are the most common North American mammals. They represent an emerging system for the genetic analyses of the physiological and behavioral bases of habitat adaptation. Phylogenetic evidence suggests a much more ancient divergence of Peromyscus from laboratory mice (Mus) and rats (Rattus) than that separating latter two. Nevertheless, early karyotypic analyses of the three groups suggest Peromyscus to be exhibit greater similarities with Rattus than with Mus. RESULTS: Comparative linkage mapping of an estimated 35% of the deer mouse genome was done with respect to the Rattus and Mus genomes. We particularly focused on regions that span synteny breakpoint regions between the rat and mouse genomes. The linkage analysis revealed the Peromyscus genome to have a higher degree of synteny and gene order conservation with the Rattus genome. CONCLUSION: These data suggest that: 1. the Rattus and Peromyscus genomes more closely represent ancestral Muroid and rodent genomes than that of Mus. 2. the high level of genome rearrangement observed in Muroid rodents is especially pronounced in Mus. 3. evolution of genome organization can operate independently of more commonly assayed measures of genetic change (e.g. SNP frequency).

Expressed sequence tags from Peromyscus testis and placenta tissue: analysis, annotation, and utility for mapping.

BACKGROUND: Mice of the genus Peromyscus are found in nearly every habitat from Alaska to Central America and from the Atlantic to the Pacific. They provide an evolutionary outgroup to the Mus/Rattus lineage and serve as an intermediary between that lineage and humans. Although Peromyscus has been studied extensively under both field and laboratory conditions, research has been limited by the lack of molecular resources. Genes associated with reproduction typically evolve rapidly and thus are excellent sources of evolutionary information. In this study we describe the generation of two cDNA libraries, one from placenta and one from testis, characterize the resulting ESTs, and describe their utility for mapping the Peromyscus genome. RESULTS: The 5' ends of 1,510 placenta and 4,798 testis clones were sequenced. Low quality sequences were removed and after clustering and contig assembly, 904 unique placenta and 2,002 unique testis sequences remained. Average lengths of placenta and testis ESTs were 711 bp and 826 bp, respectively. Approximately 82% of all ESTs were identified using the BLASTX algorithm to Mus and Rattus, and 34 - 54% of all ESTs could be assigned to a biological process gene ontology category in either Mus or Rattus. Because the Peromyscus genome organization resembles the Rattus genome more closely than Mus we examined the distribution of the Peromyscus ESTs across the rat genome finding markers on all rat chromosomes except the Y. Approximately 40% of all ESTs were specific to only one location in the Mus genome and spanned introns of an appropriate size for sequencing and SNP detection. Of the primers that were tried 54% provided useful assays for genotyping on interspecific backcross and whole-genome radiation hybrid cell panels. CONCLUSION: The 2,906 Peromyscus placenta and testis ESTs described here significantly expands the molecular resources available for the genus. These ESTs allow for specific PCR amplification and broad coverage across the genome, creating an excellent genetic marker resource for the generation of a medium-density genomic map. Thus, this resource will significantly aid research of a genus that is uniquely well-suited to both laboratory and field research.

Frequent Harderian gland adenocarcinomas in inbred white-footed mice (Peromyscus leucopus).

In 1997, three lines of inbred Peromyscus leucopus--GS109A, GS16A1, and GS16B--were acquired by the Peromyscus Genetic Stock Center. Since then, records have been kept on tumors detected by visible inspection of live animals. The inbred lines GS109A and GS16A1 presented tumors with frequencies substantially higher than that of the other inbred line or of random-bred P. leucopus stock. The average age of detection was 456 +/- 75 days (n = 24) for GS109A and 568 +/- 168 days (n = 12) for GS16A1 respectively. Surprisingly, the majority of the tumors (23 of 24 for GS109A and 8 of 12 for GS16A1) appeared to be Harderian gland lesions. During the same time period only a single tumor, a fibrosarcoma, was noted in the other inbred strain (GS16B), and one Harderian gland tumor was detected in the random bred stock. On the basis of the number of animals born to each group, tumor frequencies were approximately 22.7%, 8.3%, 0.67%, and 0.07%, for GS109A, GS16A1, GS16B, and randombred P. leucopus stock, respectively. The periocular tumors appeared to be highly malignant, with elevated mitotic indices, marked anaplasia, and metastases to regional lymph nodes and lungs. The tumors were readily transplantable to other animals of the same line. Among various other species, malignant Harderian gland tumors are relatively rare.

Distal and proximal cis-linked sequences are needed for the total expression phenotype of the mouse alcohol dehydrogenase 1 (Adh1) gene.

Mouse alcohol dehydrogenase 1 (Adh1) gene expression occurs at high levels in liver and adrenal, moderate levels in kidney and intestine, low levels in a number of other tissues, and is undetectable in thymus, spleen and brain by Northern analysis. In transgenic mice, a minigene construct containing 10 kb of upstream and 1.5 kb of downstream flanking sequence directs expression in kidney, adrenal, lung, epididymis, ovary and skin but promotes ectopic expression in thymus and spleen while failing to control expression in liver, eye, intestine and seminal vesicle. Cosmids containing either 7 kb of upstream and 21 kb of downstream or 12 kb of upstream and 23 kb of downstream sequence flanking genetically marked Adh1 additionally promotes seminal vesicle expression suggesting downstream or intragenic sequence controls expression in this tissue. However, expression in liver, adrenal, or intestine is not promoted. The Adh1(a) allele on the cosmid expresses an enzyme electrophoretically distinct from that of the endogenous Adh1(b) allele, and presence of the heterodimeric enzyme in expressing tissues confirms that transgene activity occurs in the same cell-type as the endogenous gene. Transgene expression levels promoted by cosmids were at physiologically relevant amounts and exhibited greater copy-number dependence than observed with minigenes. Transgene mRNA expression correlated with expression measured at the enzyme level. A bacterial artificial chromosome containing 110 kb of 5'- and 104 kb of 3'-flanking sequence surrounding the Adh1 gene promoted expression in tissues at levels comparable to the endogenous gene most importantly including liver, adrenal and intestinal tissue where high level Adh1 expression occurs. Transgene expression in liver was in the same cell types as promoted by the endogenous gene. Although proximal elements extending 12 kb upstream and 23 kb downstream of the Adh1 gene promote expression at physiologically relevant levels in most tissues, more distal elements are additionally required to promote normal expression levels in liver, adrenal and intestinal tissue where Adh1 is most highly expressed.

Hematologic and serum biochemical values of 4 species of Peromyscus mice and their hybrids.

Deer mice (Peromyscus maniculatus) and congeneric species are used in a wide variety of research applications, particularly studies of developmental, physiologic, and behavioral characteristics associated with habitat adaptation and speciation. Because peromyscine mice readily adapt to colony conditions, animals with traits of interest in the field are moved easily into the laboratory where they can be studied under controlled conditions. The purpose of this study was to determine the serum chemistry and hematologic parameters of 4 frequently used species from the Peromyscus Genetic Stock Center species (P. californicus, P. leucopus, P. maniculatus, and P. polionotus) and to determine quantitative differences in these parameters among species and between sexes. Triglyceride values were substantially higher in female compared with male mice in all 4 species. Similar cross-species differences in MCH were present. Overall there was considerable interspecific variation for most blood parameters, with little evidence for covariation of any 2 or more parameters. Because crosses of P. maniculatus and P. polionotus produce fertile offspring, segregation analyses can be applied to determine the genetic basis of any traits that differ between them, such as their 3.8- and 2.1-fold interspecific differences in cholesterol and triglyceride levels, respectively. The current data provide a set of baseline values useful for subsequent comparative studies of species experiencing different circumstances, whether due to natural variation or anthropogenic environmental degradation. To enable such comparisons, the raw data are downloadable from a site maintained by the Stock Center (http://ww2.biol.sc.edu/∼peromyscus).

Caring for Peromyscus spp. in research environments.

Peromyscus spp. are the most abundant native North American mammals. They have gained popularity as research animals in the last 20 years, and this trend is expected to continue as new research tools, such as whole genome sequences, baseline physiological data and others, become available. Concurrently, advances have been made in the recommendations for the care of laboratory animals. The authors provide insight into how the Peromyscus Genetic Stock Center successfully breeds and maintains several stocks of deer mice and related species. This information is beneficial to researchers that plan to include Peromyscus spp. in their research programs.

Peromyscus (deer mice) as developmental models.

Deer mice (Peromyscus) are the most common native North American mammals, and exhibit great natural genetic variation. Wild-derived stocks from a number of populations are available from the Peromyscus Genetic Stock Center (PGSC). The PGSC also houses a number of natural variants and mutants (many of which appear to differ from Mus). These include metabolic, coat-color/pattern, neurological, and other morphological variants/mutants. Nearly all these mutants are on a common genetic background, the Peromyscus maniculatus BW stock. Peromyscus are also superior behavior models in areas such as repetitive behavior and pair-bonding effects, as multiple species are monogamous. While Peromyscus development generally resembles that of Mus and Rattus, prenatal stages have not been as thoroughly studied, and there appear to be intriguing differences (e.g., longer time spent at the two-cell stage). Development is greatly perturbed in crosses between P. maniculatus (BW) and Peromyscus polionotus (PO). BW females crossed to PO males produce growth-restricted, but otherwise healthy, fertile offspring which allows for genetic analyses of the many traits that differ between these two species. PO females crossed to BW males produce overgrown but severely dysmorphic conceptuses that rarely survive to late gestation. There are likely many more uses for these animals as developmental models than we have described here. Peromyscus models can now be more fully exploited due to the emerging genetic (full linkage map), genomic (genomes of four stocks have been sequenced) and reproductive resources.

Adaptive genetic variation, stress and glucose regulation.

Elevated glucose levels in the presence of insulin are indicative of type 2 diabetes and the more inclusive metabolic syndrome. Alleles conferring susceptibility to these and other common conditions may be adaptations to past environments. It is possible that other mammals exhibiting environmental diversity harbor similar variants; therefore, we assessed glucose regulation in two species of deer mice (Peromyscus), a diverse endemic North American group. The prairie deer mouse, P. maniculatus bairdii (BW), and the Oldfield mouse, P. polionotus subgriseus (PO) differ in sexual dimorphism, behavior and habitat. PO animals exhibit better regulatory ability than BW animals, particularly among males, although both species display equivalent insulin levels/responses and non-fasted glucose levels. Hybrid males exhibit a PO glucose challenge response and subsequent analysis of consomic animals implicates Y chromosome variation as the genetic cause. Two pieces of evidence indicate that the male glucose regulatory differences are mediated by stress response: (1) fasting and handling alone account for most of the variation; (2) an inhibitor of glucocorticoid (GC) stress hormone synthesis eliminates these differences. PO males have GC levels that are twice those of BW males, indicating the presence of alleles that attenuate the GC response. We hypothesize that the interspecific physiological and behavioral differences are interrelated and that similar human variants exist.

Development of a deer mouse whole-genome radiation hybrid panel and comparative mapping of Mus chromosome 11 loci.

A 5000-rad whole-genome radiation hybrid cell panel (BW5000) was developed for mapping the deer mouse (Peromyscus maniculatus bairdii) genome. The panel consists of 103 cell lines and has an estimated marker retention frequency of 63.9% (range, 28%-88%) based on PCR typing of 30 Type I (coding gene) and 25 Type II (microsatellite) markers. Using the composite Mus map, Type I markers were selected from six Mus chromosomes, 22 of which are on Mus Chr 11. Fifteen of the Mus Chr 11 markers were simultaneously mapped on an interspecific (P. maniculatus x P. polionotus) backcross panel to test the utility of the radiation hybrid panel, create a framework map, and help establish gene order. The radiation hybrids have effectively detected linkage in the deer mouse genome between markers as far apart as 6.7 cM and resolved markers that are, in the Mus genome, as close as 0.2 Mb. Combined results from both panels have indicated a high degree of gene order conservation of the telomeric 64 cM of Mus Chr 11 in the deer mouse genome. The remaining centromeric portion also shows gene order conservation with the deer mouse but as a separate linkage group. This indicates a translocation of that portion of Mus Chr 11 in P. maniculatus and is consistent with rearrangement breakpoints observed between Mus and other mammalian genomes, including rat and human. Furthermore, this separate linkage group is likely to reside in a chromosomal region of inversion polymorphism between P. maniculatus and P. polionotus.

Inverted Gcg/CGC trinucleotide microsatellites in the 5'-region of Mus IDS mRNA: recurrent induction of aberrant reverse transcripts.

An investigation was initiated to explore previously published results indicating that approximately 80 bp of the 5'-end of the iduronate sulfatase (IDS) cDNA sequence (Accession No L07291) are 100% homologous with the 3'-UTR of isoform I of the sodium hydrogen exchanger (Acc. No. U51112). 5'-RACE carried out on IDS mRNA demonstrated the apparent homology to be a cloning artifact. A sequence comparison of the IDS 5'-RACE product with a mouse BAC clone covering the region, and with various IDS ESTs, suggested that the region is highly susceptible to cloning artifacts, a common one of which is template switching by reverse transcriptase. The nucleotide sequence flanking the translation start site is unusual in containing two inverted repeats composed of the complementary trinucleotide microsatellites, (GCG)9 and (CGC)6. These likely form a highly stable stem of 20-21 nt, through which reverse transcription is compromised. Such a stem could be involved in the regulation of IDS expression by directly affecting translation, message turnover, or serving as a substrate for siRNA production. Though such mRNA features are relatively rare, they may be more abundant but overlooked due to difficulties in their reverse transcription.

Influences of inbreeding and genetics on telomere length in mice.

We measured telomere lengths of blood leukocytes in several inbred and outbred mammalian species, using a telomere-specific fluorescent probe and flow cytometry. Humans, non-human primates, and three outbred populations of Peromyscus mice ( Peromyscus leucopus, Peromyscus maniculatus, and Peromyscus polionotus) have short telomeres. Two common strains of laboratory mice, C57BL/6J and DBA/2J, have telomeres several times longer than most other mammals surveyed. Moreover, the two inbred laboratory mouse strains display significantly different telomere lengths, suggesting the existence of strain-specific genetic determinants. To further examine the effects of inbreeding, we studied three Peromyscus leucopus inbred lines (GS109, GS16A1, and GS16B), all derived from the outbred P. leucopus stock. Telomeres of all three inbred lines are significantly lengthened relative to outbred P. leucopus, and the three lines display strain-specific significantly different telomere lengths, much like the C57BL/6J and DBA/2J strains of M. musculus. To further characterize the genetic inheritance of telomere length, we carried out several crosses to obtain hybrid F(1) mice between parental strains displaying the phenotype of long and short telomeres. In all F(1) mice assayed, peripheral blood leukocyte telomere length was intermediate to that of the parents. Additionally, we generated F(2) mice from a cross of the ( P. leucopus outbred x GS16B)F(1). Based on the distribution of telomere length in the F(2) population, we determined that more than five loci contribute to telomere length regulation in Peromyscus. We concluded that inbreeding, through unknown mechanisms, results in the elongation of telomeres, and that telomere length for a given species and/or sub-strain is genetically determined by multiple segregating loci.