Sixteen diverse laboratory mouse reference genomes define strain-specific haplotypes and novel functional loci.

We report full-length draft de novo genome assemblies for 16 widely used inbred mouse strains and find extensive strain-specific haplotype variation. We identify and characterize 2,567 regions on the current mouse reference genome exhibiting the greatest sequence diversity. These regions are enriched for genes involved in pathogen defence and immunity and exhibit enrichment of transposable elements and signatures of recent retrotransposition events. Combinations of alleles and genes unique to an individual strain are commonly observed at these loci, reflecting distinct strain phenotypes. We used these genomes to improve the mouse reference genome, resulting in the completion of 10 new gene structures. Also, 62 new coding loci were added to the reference genome annotation. These genomes identified a large, previously unannotated, gene (Efcab3-like) encoding 5,874 amino acids. Mutant Efcab3-like mice display anomalies in multiple brain regions, suggesting a possible role for this gene in the regulation of brain development.

Inheritance patterns of noise vulnerability and "protectability" in (C57BL/6J × CBA/J) F1 hybrid mice.

BACKGROUND: Interindividual variation in cochlear vulnerability to noise and ototoxins must in part reflect allelic variation in genes that largely remain unknown. Work in our laboratory has shown that young adult CBA/J mice are more vulnerable to cochlear noise injury than are similar-aged mice of other well-studied strains such as C57BL/6J (B6). Conversely, young CBA/J mice are dramatically protected against noise exposure by low-dose kanamycin (KM) treatment, while B6 mice are not. Genetic differences that distinguish these two strains may include genes that help establish the early "sensitive period" in mammals, as well as genes that shape innate protective responses to stress. These genes may have human homologs that exert similar influences and thereby partly govern individual risk of acquired hearing loss. PURPOSE: We hypothesize that young CBA/J and B6 mice carry different alleles at unknown loci that mediate their characteristic sensitivities to noise and responses to kanamycin. The first step in any experimental genetic analysis of two divergent populations is to examine F1 hybrids formed from these. Accordingly, we evaluated both noise vulnerability and the extent of protection from noise by low-dose KM in 6-wk-old F1 hybrids derived from a B6 × CBA/J cross. STUDY SAMPLE: The study included 52 CBA/J, 59 C57BL/6J (B6), and 45 (B6 × CBA/J) F1 hybrid mice, aged 6 wk at time of noise exposure. Both genders were included. INTERVENTION: For experiments aimed at noise vulnerability, B6 and F1 mice were exposed to loud broadband noise (4-45 kHz, 110 dB SPL) for varying durations, and the resulting noise-induced permanent threshold shifts (NIPTSs, measured 2 wk postnoise) were compared with previous data from CBA/J mice. For experiments aimed at KM-based "protectability," CBA/J, B6, and F1 mice received either kanamycin (300 mg/kg, sc) or saline twice daily for 10 days and then were noise exposed for 30 min, followed by measurement of NIPTS at 2 wk postnoise. DATA COLLECTION AND ANALYSIS: Data comprised auditory brainstem response (ABR) thresholds examined by two-way ANOVA (threshold × frequency, group) and derived metrics for NIPTS, plotted versus noise duration. RESULTS: The "threshold" noise exposure duration for NIPTS in F1 hybrid mice was similar to that in CBA/J. Like CBA/J mice, F1 mice were also significantly protected from noise by KM although the protection appeared less robust than in the CBA/J parent strain. B6 mice appeared harmed by KM alone, even without noise exposure. None of the experimental groups provided any evidence for synergistic interactions between noise and KM. CONCLUSIONS: Our data support the hypothesis that young CBA/J and B6 mice carry different alleles that underlie their divergent responses to KM and sensitivities to noise exposure. While the number and type of genes remain unknown, they are worth pursuing because they establish completely novel hearing phenotypes with potential relevance to humans. Our results lay the foundation for mapping of the underlying genes, and ultimately gene identification.

Gravity receptor aging in the CBA/CaJ strain: a comparison to auditory aging.

The CBA/CaJ mouse strain is commonly used as a control as it has no known genetic mutations affecting the inner ear, maintains hearing sensitivity throughout life, and serves as a background for creating new genetic strains. The purpose of the present study was to characterize the effects of age and gender on gravity receptor function and compare functional changes between auditory and vestibular modalities. Vestibular-evoked potentials (VsEPs), auditory brainstem responses (ABRs), and distortion product otoacoustic emissions were measured in 131 mice. VsEP thresholds deteriorated an average of 0.39 dB re: 1.0 g/ms per month and at the oldest ages (18-23 months old) showed an average loss of 49% of VsEP dynamic range. No significant gender differences were found for VsEPs. ABR thresholds increased by an average of 1.35, 1.38, and 1.15 dB pe SPL per month for ABR stimulus frequencies of 8, 16, and 32 kHz, respectively, demonstrating an average decrease in auditory dynamic range of 25-35% at advanced ages. Both modalities declined with age. Age-related decreases in gravity receptor sensitivity should be considered when using the CBA/CaJ strain for vestibular studies.

Sperm mitochondria diaphorase activity--a gene mapping study of recombinant inbred strains of mice.

In order to study the genetic control of semen quality parameters, we derived a set of recombinant inbred (RI) mice from crosses between two inbred strains, KE and CBA/Kw, which differ significantly in gamete quality and fertility parameters. In this work, we used male mice from the two parental strains and from ten RI strains to map genes controlling quantitative traits such as sperm mitochondrial diaphorase activity, and assessed the correlation between this trait, sperm motility and in vivo fertilization efficiency. We analyzed sperm mitochondrial dehydrogenase (diaphorase) activity (NADH-dependent NBT assay) cytochemically by means of computerized image densitometry and obtained values for four parameters: 1) integrated optical density (IOD) for all pixels of the midpiece, 2) mean optical density (MOD) for the midpiece pixels, 3) length of sperm midpiece and 4) area of sperm midpiece. Polymorphic microsatellite marker profiles were prepared for 20 mouse chromosomes in the ten RI strains. We used Map Manager QTX software to correlate the strain distribution patterns (SDPs) of the four measured parameters with the SDPs of the analyzed markers. Hypothetical genes modifying diaphorase activity were mapped to chromosomal region 19q43-19q47, containing, for example, Poll, Sfxn2, Cyp17a1 and Usmg5 genes. Chromosomal regions 18q44 and 18q49-18q80 also showed correlation with the SDPs of diaphorase activity. Katnal2, Me2 and StARD6 candidate genes were proposed from this region. Diaphorase activity in the mouse sperm midpiece did not correlate with in vivo fertilization efficiency, but was negatively correlated with the linearity and straightness of sperm movement.

Variation of the response to the optokinetic drum among various strains of mice.

The optokinetic drum has become an appropriate tool to examine visual properties of mice. We performed baseline measurements using mice of the inbred strains C3H, C57BL/6, BALB/c, JF1, 129 and DBA/2 at the age of 8-15 weeks. Each individual C57BL/6, 129 and JF1 mouse was reliably identified as non-affected in vision by determining head-tracking responses. C3H mice were used as negative control because of their inherited retinal degeneration; as expected, they did not respond to the moving stripe pattern. Surprisingly, BALB/c and DBA/2 mice showed the same result. Electroretinography, funduscopy and histology of BALB/c mice did not reveal any abnormality concerning the structure or function of the retina and the remaining eye. Therefore, it might be assumed that BALB/c mice suffer from disturbances of the central visual system. Preliminary results from linkage analysis of the non-responding phenotype in the BALB/c mice indicate a recessive, monogenic mode of inheritance; the causative gene is located on chromosome 7, but significantly different from the albino locus. In conclusion, C57BL/6, 129 and JF1 represent appropriate inbred strains for high throughput screenings with the optokinetic drum.

Individual and typological behavioral characteristics of CBA/CaLac mice.

Individual characteristics of higher nervous activity were studied in CBA/CaLac mice. The animals were divided into groups by the parameters of drinking response conditioning in a complex spatial environment. As distinct from bad learners, good learners exhibited higher orientation and exploratory activity in the open field, rapid adaptation, and low ability to retain the responses. Changes in conditioned reflex activity during neuroses were more pronounced in good learners.

Genetic linkage analysis of pulmonary fibrotic response to silica in mice.

Inter-individual variations in the development of silicosis, even within the same environments, have been reported, which suggest the contribution of genetic factors in silicosis aetiology. The aim of the present study was to determine whether there is any significant genetic influence on the development of silicosis. Furthermore, which genetic loci are responsible for the pulmonary response to silica exposure? Eight strains of inbred mice were used to examine the genetic influence on the lung fibrotic response to silica exposure. After intercross-breeding between the most susceptible and most resistant strains, a genome-wide linkage analysis of quantitative trait loci (QTL) was performed. Hydroxyproline was applied as an index, and genotypes of 167 marker genes were analysed by fragment analysis using a capillary-type sequencer. There was significant inter-strain difference in the mean concentration of hydroxyproline contents among the eight strains of mice. Breeding studies were conducted between the most susceptible, C57BL/6J, and the most resistant strain, CBA/J. A genome-wide linkage analysis of silica-exposed intercrossed cohorts identified significant QTL on chromosome 4 and suggestive QTL on chromosomes 3 and 18. The present study demonstrates that genetic factors may play a significant role in fibrotic-lung responses to silica; one significant and two suggestive quantitative trait loci were identified.

Gene expression variation between mouse inbred strains.

BACKGROUND: In this study, we investigated the effect of genetic background on expression profiles. We analysed the transcriptome of mouse hindlimb muscle of five frequently used mouse inbred strains using spotted oligonucleotide microarrays. RESULTS: Through ANOVA analysis with a false discovery rate of 10%, we show that 1.4% of the analysed genes is significantly differentially expressed between these mouse strains. Differential expression of several of these genes has been confirmed by quantitative RT-PCR. The number of genes affected by genetic background is approximately ten-fold lower than the number of differentially expressed genes caused by a dystrophic genetic defect. CONCLUSIONS: We conclude that evaluation of the effect of background on gene expression profiles in the tissue under study is an effective and sensible approach when comparing expression patterns in animal models with heterogeneous genetic backgrounds. Genes affected by the genetic background can be excluded in subsequent analyses of the disease-related changes in expression profiles. This is often a more effective strategy than backcrossing and inbreeding to obtain isogenic backgrounds.

The development of a highly informative mouse Simple Sequence Length Polymorphism (SSLP) marker set and construction of a mouse family tree using parsimony analysis.

To identify highly informative markers for a large number of commonly employed murine crosses, we selected a subset of the extant mouse simple sequence length polymorphism (SSLP) marker set for further development. Primer pairs for 314 SSLP markers were designed and typed against 54 inbred mouse strains. We designed new PCR primer sequences for the markers selected for multiplexing using the fluorescent dyes FAM, VIC, NED, and ROX. The number of informative markers for C57BL/6J x DBA/2J is 217, with an average spacing of 6.8 centiMorgans (cM). For all other pairs of strains, the mean number of informative markers per cross is 197.0 (SD 37.8) with a mean distance between markers of 6.8 cM (SD 1.1). To confirm map positions of the 224 markers in our set that are polymorphic between Mus musculus and Mus spretus, we used The Jackson Laboratory (TJL) interspecific backcross mapping panel (TJL BSS); 168 (75%) of these markers had not been previously mapped in this cross by other investigators, adding new information to this community map resource. With this large data set, we sought to reconstruct a phylogenetic history of the laboratory mouse using Wagner parsimony analysis. Our results are largely congruent with the known history of inbred mouse strains.

[Genetic and molecular characteristics of a new natural haplotype twMP1 of the house mouse (Mus domesticus R.) from Peru]. / Geneticheskaia i molekuliarnaia kharakteristika novogo prirodnogo gaplotipa twMP1 domovoi myshi (Mus domesticus) iz Peru.

A new natural haplotype, twWP1, found in a population of house mouse Mus domesticus from Peru, was subjected to genetic and molecular analyses. Experiments were performed to study the complementation of the new haplotype, fertility of twMP1/tx heterozygotes, and transmission ratio distortion (TRD) of the t-carrying chromosome in the progeny of heterozygous males. Molecular analysis included blot hybridization with t-specific probes Tu48, Tu66, and Tu119. The results were collated with the structure and properties of the t complex, and the new haplotype was identified as a complete lethal one.

Identification and functional characterization of novel polymorphisms associated with the genes for arylamine N-acetyltransferases in mice.

Arylamine N-acetyltransferase (NAT) polymorphism in humans has been associated with variation in susceptibility to drug toxicity and cancer. In mice, three NAT isoenzymes are encoded by Nat1, Nat2 and Nat3 genes. Only Nat2 has been shown previously to be polymorphic, a single nucleotide substitution causing the slow acetylator phenotype in the A/J strain. We sequenced the Nat genes from inbred (CBA and 129/Ola), outbred (PO and TO) and wild-derived inbred (Mus spretus and Mus musculus castaneus) mouse strains and report polymorphism in all three Nat genes of M. spretus and in Nat2 and Nat3 genes of M. m. castaneus. Enzymatic activity assays using liver homogenates demonstrated that M. m. castaneus is a 'fast' and M. spretus a 'slow' acetylator. Western blot analysis indicated that hepatic NAT2 protein is less abundant in M. spretus than M. m. castaneus. The new allozymes were expressed in a mammalian cell line and NAT enzymatic activity was measured with a series of substrates. NAT1 and NAT2 isoenzymes of M. m. castaneus exhibited a higher rate of acetylation, compared with those of M. spretus. Activity of the NAT3 allozymes was hardly detectable, although the Nat3 gene does appear to be transcribed, since mRNA was detected by RT-PCR in the spleen. Additional polymorphisms, useful for Nat-related genetic studies, have been identified between BALB/c, C57Bl/6J, A/J, 129/Ola, CBA, PO, TO, M. m. castaneus and M. spretus strains in four microsatellite repeats located close to the Nat genes.

Virulence in rodent malaria: host genotype by parasite genotype interactions.

In an effort to understand what limits the virulence of malaria parasites, we infected inbred mice of three genotypes (C57Bl/6J, CBA/Ca and DBA/2) with one of two parasite lines of the rodent malaria Plasmodium chabaudi. One of these parasite lines had been serially passaged through C57Bl/6J mice and had evolved higher asexual growth rate, virulence and transmission in the process. The other parasite line was the unadapted ancestral line which had low virulence. In all three host genotypes, the C57Bl/6J-adapted parasite line was more virulent than the ancestral line thus indicating that trade-offs in virulence between alternative host genotypes had not placed strong constraints on the evolution of high virulence in this system. By examining the infection dynamics for fitness-related components-asexual parasite population growth, transmission and virulence-we revealed alternative possible explanations for what sets the upper limit to virulence in nature. The total number of transmission forms (gametocytes) produced during the infection, a measure of parasite Darwinian fitness, was four-fold higher in mice that survived the infection than those which died. Among mice that survived, total gametocyte production was greatest in the host genotype that suffered intermediate levels of morbidity (anaemia and weight loss). Thus, there were transmission costs of high virulence that were partly due to host death (as most theoretical models of virulence evolution assume), but perhaps partly due to some factor related to high morbidity. Both mortality and morbidity-related factors might therefore influence the upper limit on virulence of malaria parasites.

Interstrain variation in murine aerobic capacity.

PURPOSE: The contribution of genetic factors to aerobic capacity is unknown. The purpose of this study was to measure maximal aerobic performance among inbred strains of mice to provide basic heritability estimates. METHODS: Eight female mice, 8 to 10 wk old, in 10 inbred strains (A/J, AKR/J, Balb/cJ, C(3)H/HeJ, C57Bl/6J, C57L/J, C(3)Heb/FeJ, CBA/J, DBA/2J, and SWR/J) were run on a treadmill until exhaustion. The protocol started at 22 m.min(-1) and increased in speed approximately 6 m.min(-1) every 4 min. After 4 min at 42.4 m.min(-1), the grade was increased 2% every 4 min thereafter until the mouse could not run off of the shock grid (150 V; 1.5 mA). RESULTS: There were significant differences between inbred strains in maximal duration of exercise accomplished (P < 0.0001). The order of strain-specific exercise duration was Balb/cJ > SWR/J > CBA/J > C57L/J > C3H/HeJ > C3Heb/FeJ > C57Bl/6J > AKR/J > DBA/2J > A/J. Two measures of heritability in the broad sense, intraclass correlation (0.73), and the coefficient of genetic determination (0.58) were both significant. CONCLUSION: These data indicate that there is a strong genetic contribution to aerobic capacity in mice.

Skin allograft maintenance in a new synchimeric model system of tolerance.

Treatment of mice with a single donor-specific transfusion plus a brief course of anti-CD154 mAb uniformly induces donor-specific transplantation tolerance characterized by the deletion of alloreactive CD8+ T cells. Survival of islet allografts in treated mice is permanent, but skin grafts eventually fail unless recipients are thymectomized. To analyze the mechanisms underlying tolerance induction, maintenance, and failure in euthymic mice we created a new analytical system based on allo-TCR-transgenic hemopoietic chimeric graft recipients. Chimeras were CBA (H-2(k)) mice engrafted with small numbers of syngeneic TCR-transgenic KB5 bone marrow cells. These mice subsequently circulated a self-renewing trace population of anti-H-2(b)-alloreactive CD8+ T cells maturing in a normal microenvironment. With this system, we studied the maintenance of H-2(b) allografts in tolerized mice. We documented that alloreactive CD8+ T cells deleted during tolerance induction slowly returned toward pretreatment levels. Skin allograft rejection in this system occurred in the context of 1) increasing numbers of alloreactive CD8+ cells; 2) a decline in anti-CD154 mAb concentration to levels too low to inhibit costimulatory functions; and 3) activation of the alloreactive CD8+ T cells during graft rejection following deliberate depletion of regulatory CD4+ T cells. Rejection of healed-in allografts in tolerized mice appears to be a dynamic process dependent on the level of residual costimulation blockade, CD4+ regulatory cells, and activated alloreactive CD8+ thymic emigrants that have repopulated the periphery after tolerization.

[Different influences of genomic imprinting on the development of parthenogenetic cell clones in C57BL/6 and CBA mice]. / Razlichnoe vliianie genomnogo imprintinga na razvitie partenogeneticheskikh kletochnykh klonov myshei C57BL/6 i CBA.

Clonal analysis of parthenogenetic chimeric mouse embryos C57B1/6(PG)<-->BALB/c has shown that parthenogenetic cell clones C57BL/6 are present in the brain, liver, and kidneys of 14- and 18-day-old embryos. The content of the parthenogenetic component (PG) in these organs on day 18 was lower than on day 14, and, in some 18-day-old embryos, parthenogenetic cell clones were absent from the liver and/or kidneys. These data suggest that, during the embryogenesis of parthenogenetic chimeras, parthenogenetic cell clones of mostly endodermal and mesodermal origins were actively eliminated. Therefore, in such parthenogenetic adult chimeras, parthenogenetic clones of mostly ectodermal origins were preserved. In parthenogenetic chimeras CBA(PG)<-->BALB/c, parthenogenetic cell clones were actively eliminated at early embryonic stages, and, as a result, they were absent at the post-implantation stages. Hence, during development of parthenogenetic cell clones, the effects of genomic imprinting are expressed unequally in C57BL/6 and CBA mice.

Quantitative trait loci affecting the behavior of A/J and CBA/J intercross mice in the elevated plus maze.

How allelic diversity affects neural mechanisms to produce behavioral variation is largely unknown. The elevated plus maze, consisting of open and closed arms, has been used as a model of behavioral variation in rodent exploration. Under dim illumination the nature of the sensory stimuli that influence arm choice is uncertain. Two inbred mouse strains, A/J (Tyrc/Tyrc, the albino phenotype, mutation in tyrosinase) with a strong preference for closed arm entry, and CBA/J (Pdebrdl/Pdebrdl, the retinal degeneration phenotype, mutation in the beta-subunit of rod cGMP phosphodiesterase), with a weak preference for open arm entry, were studied under varying light. Because behavioral differences persist under red light, variation in light perception is not likely to fully account for variation in arm choice. To identify genetic factors influencing arm choice (100 x Open arm entries/Total arm entries) quantitative trait loci analyses (QTL) were performed on (A/J x CBA/J)F2 mice. Two QTLs, one of which includes PDEB, were identified on Chr 5 (LOD > 10) and account for > 30% of the behavioral variation in arm preference. Tyr (Chr 7, 44 cM) was linked to closed arm entries but not arm preference, and is unlikely to be acting through a direct effect on light perception, because A/J arm entries were not affected by red light and there was no interaction with PDEB in the (A/J x CBA/J)F2 mice. Whether the candidate QTLs on Chr 5 affect arm choice through an effect on light perception is unknown, but phenotypic differences between F2 mice with retinal degeneration and CBA/J mice and F2 mice with albinism and A/J mice suggest that factors other than light sensitivity contribute to arm preference in these two strains.

Features of the genetically defined anxiety in mice.

The behaviors of male mice of the C57BL/6J (C57), CBA/Lac (CBA) and BALB/c (BALB) strains have been studied in the plus-maze and open field tests for estimation of state anxiety in the stressful novel conditions, and in the cubic box test (exploration of novel cubic box) and the partition test (behavioral reactivity to the unfamiliar partner in the neighboring compartment) for estimation of trait anxiety in the unstressful familiar conditions of the home cage. Plus-maze data suggest that C57 mice are the more anxious than CBA and BALB ones. However, it was revealed the opposite rank order in the open field. The study on the effect of pre-testing in the one of test on the behavior in the other test revealed active behavioral strategy in C57 mice in any situations. The plus-maze behavior of CBA mice was affected to a much lesser extent than in C57 ones after pre-testing in the open field, but expressed changes were observed in open field behavior of CBA mice after pre-testing in the plus-maze. BALB mice displayed low-reactive behavior after any pre-testing exposure under the state anxiety-provoking conditions. Familiar environment revealed a higher level of trait anxiety in C57 than males of other two strains: CBA and BALB mice willingly explore unfamiliar partner and cubic box while C57 mice avoid its. Mainly genetically inherent state anxiety in CBA mice and trait anxiety in C57 mice has been suggested. Lowest state and trait indices of anxiety were revealed in BALB mice in these conditions.

High resolution magnetic resonance angiography non-invasively reveals mouse strain differences in the cerebrovascular anatomy in vivo.

High resolution magnetic resonance angiography (MRA) revealed highly variable arterial cerebrovascular structures in mice from different strains and within the same strain. C57Black/6 mice presented small unilateral anastomoses between the posterior cerebral and the superior cerebellar arteries. Well developed, either unilateral or bilateral, posterior communicating arteries (PcomA) were detected on CBA mice. The arterial structure of CD1 mice ranged from no detectable anastomoses to well developed, unilateral PcomAs. SV-129 mice showed significantly shorter middle cerebral arteries compared to the other strains, and clear bilateral anastomoses between the posterior cerebral and the superior cerebellar arteries. Because of its non-invasiveness, MRA may be of importance in murine stroke studies by enabling the selection of animals and/or the side for performing the surgical intervention, and the verification of its success. Magn Reson Med 44:252-258, 2000.

Quantitative measure of genetic differences in susceptibility to noise-induced hearing loss in two strains of mice.

The CBA/CaJ (CB) and C57BL/6J (B6) inbred strains of mice were exposed for 1 h to noise intensities between 98 and 119 dB SPL. Previous studies indicated that the B6 mice exhibited permanent threshold shifts (PTS) after 1h exposure to 110 dB, whereas the CB mice did not exhibit any PTS. These differences in susceptibility to noise-induced hearing loss (NIHL) appear to be due to a gene for age-related hearing loss (AHL). The current study was designed to determine dose-response curves for NIHL over the ranges of intensities of noise that would characterize the B6 and CB inbred strains of mice. Because of the considerable differences in sensitivity to NIHL, the noise exposures for the two strains overlapped only at 110 and 113 dB. Nevertheless, the two strains exhibited two different dose-response curves, offset and with different slopes. We postulate that the B6 strain of mice exhibits a more linear increase for PTS from 98-113 dB, consistent with incremental effects on some metabolic physiological mechanism(s); the abrupt transition in NIHL between 113 and 116 dB for the CB mice is consistent with an ototraumatic structural injury.

[The characteristics of the manifestation of hereditarily induced anxiety in male C57Bl/6J and CBA/Lac mice]. / Osobennosti proiavleniia nasledstvenno-obuslovlennoi trevozhnosti u samtsov myshei linii C57Bl/6J i CBA/Lac.

Behavior of male mice of C57Bl/6J and CBA/Lac strains was tested in the elevated plus-maze and open field in order to estimate state anxiety in novel conditions. The cube and partition tests were used to reveal trait anxiety in the familiar conditions of the home cage. It is concluded that genetically defined state anxiety is more pronounced in CBA/Lac mice and trait anxiety in C57Bl/6J strain.