Detected microsatellite polymorphisms in genetically altered inbred mouse strains

Microsatellites are 50–200 repetitive DNA sequences composed of 1- to 6-base-pair-long reiterative motifs within the genome. They are vulnerable to DNA modifications, such as recombination and/or integration, and are recognized as “sentinel” DNA. Our previous report indicated that the genotypes of the microsatellite loci could change from mono- to poly-morphisms (CMP) in gene knockout (KO) mice, implying that genetic modification induces microsatellite mutation. However, it is still unclear whether the random insertion of DNA fragments into mice genomes produced via transgene (Tg) or N-ethyl-N-nitrosourea (ENU) would also result in microsatellite mutations or microsatellite loci genotypes changes. This study was designed to find possible clues to answer this question. In brief, 198 microsatellite loci that were distributed among almost all of the chromosomes (except for the Y) were examined through polymerase chain reaction to screen possible CMPs in six Tg strains. First, for each strain, the microsatellite sequences of all loci were compared between Tg and the corresponding background strain to exclude genetic interference. Simultaneously, to exclude spontaneous mutation-related CMPs that might exist in the examined six strains, mice from five spontaneously mutated inbred strains were used as the negative controls. Additionally, the sequences of all loci in these spontaneous mutated mice were compared to corresponding genetic background controls. The results showed that 40 of the 198 (20.2 %) loci were identified as having CMPs in the examined Tg mice strains. The CMP genotypes were either homozygous or heterozygous compared to the background controls. Next, we applied the 40 CMP positive loci in ENU-mutated mice and their corresponding background controls. After that, a general comparison of CMPs that exist among Tg, ENU-treated and KO mouse strains was performed. The results indicated that four (D11mit258, D13mit3, D14mit102 and DXmit172) of the 40 (10 %) CMP loci were shared by Tg and KO mice, two (D15mit5 and D14mit102) (5 %) by Tg and ENU-treated mice, and one (D14mit102) (2.5 %) by all three genetic modifications. Collectively, our study implies that genetic modifications by KO, Tg or chemical mutant can trigger microsatellite CMPs in inbred mouse strains. These shared microsatellite loci could be regarded as “hot spots” of microsatellite mutation for genetic monitoring in genetic modified mice.     

Gene markers for alcohol-metabolizing enzymes among recombinant inbred strains of mice with differential behavioural responses towards alcohol

The genetic variability of alcohol dehydrogenase (C2 isozyme), aldehyde dehydrogenase (A2 isozyme) and aldehyde oxidase (A2 isozyme) has been examined among recombinant inbred strains of mice which have been previously studied concerning their differential behavioural responses towards alcohol. The results showed no correlation between biochemical phenotype for these loci and behavioural response.     

Gene markers for alcohol-metabolizing enzymes among recombinant inbred strains of mice with differential behavioural responses towards alcohol

The genetic variability of alcohol dehydrogenase (C₂ isozyme), aldehyde dehydrogenase (A₂ isozyme) and aldehyde oxidase (A₂ isozyme) has been examined among recombinant inbred strains of mice which have been previously studied concerning their differential behavioural responses towards alcohol. The results showed no correlation between biochemical phenotype for these loci and behavioural response.     

Establishment and characteristics of five analbuminemic inbred strains of rats

Five analbuminemic inbred strains of rats (AD/1, AD/2, AD/3, AD/4, AD/5) were established from Nagase analbuminemic rats (NAR). They showed no genetic differences in coat color, biochemical marker gene loci and skin grafting test. Their serum levels of total cholesterol, phospholipids, triglycerides, and beta-lipoproteins were compared with normal inbred strains (L) derived from Sprague-Dawley rats. Their plasma apoproteins were also examined. All inbred strains of analbuminemic rats showed hyperlipidemia progressing with age although there were slight variations in their lipid and apoprotein levels. These analbuminemic inbred strains of rats may be multigenic models of lipid metabolism abnormality.     

Allelic profile at 37 biochemical loci of two inbred strains of the house mouse derived from wild Mus musculus musculus

Two newly established inbred strains derived from Mus musculus musculus, designated PWD/Ph (F29) and PWK/Ph (F33), were examined for their alleles at 37 biochemical loci located on 12 different chromosomes. The allelic pattern showed characteristic differences from those observed in common inbred strains. The genetic distance D between PWK/Ph and PWD/Ph was 0.027, whereas the corresponding values for the genetic distances between PWK/Ph and C57BL/6J, DBA/2J, BALB/cJ and SWR/J were 0.777, 0.721, 0.721 and 0.838 respectively. New allozymes are described as being controlled by the loci Es-23, Pre-2 and Tam-1. The genetic relationship to M.m.molossinus is indicated by identical alleles at six other loci (Es-2, Es-9, Es-10, Es-11, Es-18 and Es-22).     

Detection of loci for allergic asthma using SMXA recombinant inbred strains of mice

Asthma is regarded as a multifactorial inflammatory disorder arising as a result of inappropriate immune responses in genetically susceptible individuals to common environmental antigens. However, the precise molecular basis is unknown. To identify genes for susceptibility to three asthma-related traits, airway hyperresponsiveness (AHR), eosinophil infiltration, and allergen-specific serum IgE levels, we conducted a genetic analysis using SMXA recombinant inbred (RI) strains of mice. Quantitative trait locus analysis detected a significant locus for AHR on chromosome 17. For eosinophil infiltration, significant loci were detected on chromosomes 9 and 16. Although we could not detect any significant loci for allergen-specific serum IgE, analysis of consomic strains showed that chromosomes 17 and 19 carried genes that affected this trait. We detected genetic susceptibility loci that separately regulated the three asthma-related phenotypes. Our results suggested that different genetic mechanisms regulate these asthma-related phenotypes. Genetic analyses using murine RI and consomic strains enhance understanding of the molecular mechanisms of asthma in human.     

Biochemical genetic variants in mice selectively bred for sensitivity or resistance to ethanol-induced sedation

The distribution of biochemical genetic variants was examined among eight inbred strains of mice, which served as contributors to a heterogeneous stock of mice (HS), and in short-sleep (SS) and long-sleep (LS) mice, selectively bred from the HS stock for differential ethanol sensitivity. Fifteen loci for enzymes of alcohol and aldehyde metabolism, as well as 12 other biochemical loci, were investigated. Thirteen of these loci exhibited allelic variation between strains, of which six were separately fixed in the SS and LS mice. Comparisons of genetic similarity coefficients, based upon the distributions of allelic variants for the loci examined, with behavioural sensitivities (sleep-time) to an acute dose of ethanol for the inbred and selected strains of mice, indicated no correlations between these data. This suggests that this collective group of loci are not useful indicators of the genes selectively bred in the SS and LS strains, which are responsible for the differential sensitivities to acute doses of ethanol.     

Genetic affinities of inbred mouse strains of uncertain origin

Phylogenetic analyses of genetic data arising from 144 gene loci are used to describe the interrelationships among 24 widely used inbred strains of mice. An unordered-parsimony analysis gives a cladogram that is virtually identical to the known genealogy of the mouse strains. A loss-parsimony analysis is used to evaluate the hypothesis that the observed patterns of genetic divergence among these 24 strains can be explained by the segregation of residual heterozygosity arising from a small population of highly heterozygous mice. The loss-parsimony cladogram is very similar to both the unordered-parsimony cladogram and the known genealogy of the mice. The phylogenetic analyses of these 144 loci are integrated with data on the type and origin of the Y chromosome. Inclusion of the Y-chromosome data provides additional insights into the genetic composition of several of the original stocks used to produce the current inbred strains of mice. Ten strains of uncertain origin are contained in these analyses, including AKR, BUB, CE, I, NZB, P, RF, SJL, ST, and SWR. SJL is hypothesized to have been derived from the same Swiss albino stock previously used to produce SWR. The BUB strain appears to have had a complex origin and shows closest genetic similarity to SWR and ST. AKR and RF are shown to be closely related, while the I strain shows greatest genetic similarity to DBA/2 for the 144 loci. However, I and DBA possess different types of Y chromosome. The NZB strain shows genetic similarity to several stocks of both U.S. and European origins. The power of the genetic data used in these analyses reiterates that inbred strains of mice can be a valuable paradigm for studies in evolutionary biology.      

Numbers and avidity of anti-DNP antibody plaques in different inbred mouse strains

Spleen cells of mice from eight inbred strains and three F₁ hybrids, undergoing a secondary immune response to dinitrophenylated keyhole limpet hemocyanin (DNP-KLH), were examined for numbers of indirect DNP-specific plaque forming cells (PFC) as well as avidity of anti-DNP antibodies. The results indicated that the magnitude of the immune response is under genetic control. Differences in average avidity and heterogeneity of avidity were found among different mouse strains, suggesting genetic control of these parameters. However, no simple pattern of inheritance for these characteristics emerged from the study.     

An investigation of genetic variation within a series of congenic strains of mice

2 congenic strains of mice, B6N.AKN-Ahk and D2N.B6N-Ahb, imported from the USA, were found to be either segregating or fixed for an incorrect allele at a number of biochemical loci. B6N.AKN-Ahk, supposedly congenic with C57BL/6N, had the wrong genotype at 6 out of 12 biochemical loci; D2N.B6N-Ahb, supposedly congenic with DBA/2N, was segregating at 3 out of 9 loci. There was genetic variation in mandible shape within the 2 strains but no abnormal coat colours were found and no hybrid vigour in breeding performance was detected. Analyses in the USA confirmed these results and showed that 2 other congenic strains, C3N.D2N-Ahd and AKN.B6J-Ahb, were also segregating at a number of loci. Some of the alleles found in the C3N.D2N-Ahd mice must be the result of a genetic contamination. The simplest explanation for this breakdown in the backcrossing programme is genetic contamination with other congenic strains or recombinant inbred lines under development in the same laboratory. These findings emphasize the importance of continual genetic monitoring of all genetic stocks at regular intervals and in particular during the development of congenic and recombinant lines.     

Nucleotide variation in wild and inbred mice

The house mouse is a well-established model organism, particularly for studying the genetics of complex traits. However, most studies of mice use classical inbred strains, whose genomes derive from multiple species. Relatively little is known about the distribution of genetic variation among these species or how variation among strains relates to variation in the wild. We sequenced intronic regions of five X-linked loci in large samples of wild Mus domesticus and M. musculus, and we found low levels of nucleotide diversity in both species. We compared these data to published data from short portions of six X-linked and 18 autosomal loci in wild mice. We estimate that M. domesticus and M. musculus diverged <500,000 years ago. Consistent with this recent divergence, some gene genealogies were reciprocally monophyletic between these species, while others were paraphyletic or polyphyletic. In general, the X chromosome was more differentiated than the autosomes. We resequenced classical inbred strains for all 29 loci and found that inbred strains contain only a small amount of the genetic variation seen in wild mice. Notably, the X chromosome contains proportionately less variation among inbred strains than do the autosomes. Moreover, variation among inbred strains derives from differences between species as well as from differences within species, and these proportions differ in different genomic regions. Wild mice thus provide a reservoir of additional genetic variation that may be useful for mapping studies. Together these results suggest that wild mice will be a valuable complement to laboratory strains for studying the genetics of complex traits.     

A Linkage Map of Endogenous Murine Leukemia Proviruses

Thirty endogenous proviruses belonging to the modified polytropic (Mpmv) class of murine leukemia virus (MLV) were identified by proviral-cellular DNA junction fragment segregation in several sets of recombinant inbred mice. Twenty-six Mpmv loci were mapped to chromosomal regions by matching proviral strain distribution patterns to those of previously assigned genes. Like other endogenous nonecotropic MLVs, Mpmv loci were present on several chromosomes in all strains examined. We pooled recombinant inbred strain linkage data from 110 MLV loci and selected marker genes in order to construct a chromosomal linkage map. Every mouse chromosome was found to harbor at least one proviral insertion, and several regions contained multiple integrations. However, the overall distribution of the 110 mapped proviruses did not deviate significantly from a random distribution. Because of their polymorphism in inbred strains of mice, and the ability to score as many as 57 proviruses per strain using only three hybridization probes, the nonecotropic MLVs mapped in common strains of mice offer a significant advantage over older methods (e.g., biochemical or individual restriction fragment polymorphisms) as genetic markers. These endogenous insertion elements should also be useful for assessing strain purity, and for studying the relatedness of common and not-so-common inbred strains.     

Origins of mouse inbred strains deduced from whole-genome scanning by polymorphic microsatellite loci

Microsatellite loci are uniformly distributed at approximately 100-kbp intervals on all chromosomes except the chromosome Y, and genetic information about more than 9000 loci and high-throughput polymorphism analysis are now available. Taking advantage of these properties, we carried out whole-genome scanning using eight common inbred strains (CIS) of laboratory mice, including A/J, C57BL/6J, CBA/J, DBA/2J, SM/J, SWR/J, NC/Nga, and 129/SvJ, and eight wild-derived inbred strains (WIS), BGL2/Ms, CAST/Ei, JF1/Ms, MSM/Ms, NJL/Ms, PGN2/Ms, SK/CamEi, and SWN/Ms. We selected and located 1226 informative loci at 1.2-cM average intervals on all of the chromosomes of the 16 strains and compared the polymorphisms of the eight CIS with those from the eight WIS as subspecies representatives. More than 50% of the loci can be identified as WIS (therefore, subspecies-specific) alleles in the CIS genomes. We also discovered that the CIS chromosomes form a mosaic structure with an average ratio of domesticus to non-domesticus alleles of 3:1. Furthermore, the domesticus alleles were present much more frequently on the CIS chromosome X than on their autosomes, suggesting that successive backcrossing of non-domesticus stocks to domesticus stocks had been undergone at the beginning of CIS history.     

A comprehensive SNP-based genetic analysis of inbred mouse strains

Dense genetic maps of mammalian genomes facilitate a variety of biological studies including the mapping of polygenic traits, positional cloning of monogenic traits, mapping of quantitative or qualitative trait loci, marker association, allelic imbalance, speed congenic construction, and evolutionary or phylogenetic comparison. In particular, single nucleotide polymorphisms (SNPs) have proved useful because of their abundance and compatibility with multiple high-throughput technology platforms. SNP genotyping is especially suited for the genetic analysis of model organisms such as the mouse because biallelic markers remain fully informative when used to characterize crosses between inbred strains. Here we report the mapping and genotyping of 673 SNPs (including 519 novel SNPs) in 55 of the most commonly used mouse strains. These data have allowed us to construct a phylogenetic tree that correlates and expands known genealogical relationships and clarifies the origin of strains previously having an uncertain ancestry. All 55 inbred strains are distinguishable genetically using this SNP panel. Our data reveal an uneven SNP distribution consistent with a mosaic pattern of inheritance and provide some insight into the changing dynamics of the physical architecture of the genome. Furthermore, these data represent a valuable resource for the selection of markers and the design of experiments that require the genetic distinction of any pair of mouse inbred strains such as the generation of congenic mice, positional cloning, and the mapping of quantitative or qualitative trait loci.The content of this publication does not necessarily reflect the view or policies of the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement by the U.S. Government.     

Analysis of microsatellite polymorphism in inbred knockout mice

Previously, we found that the genotype of 42 out of 198 mouse microsatellite loci, which are distributed among all chromosomes except the Y chromosome, changed from monomorphism to polymorphism (CMP) in a genetically modified inbred mouse strain. In this study, we further examined whether CMP also relates to the homologous recombination in gene knockout (KO) mouse strains. The same 42 microsatellite loci were analyzed by polymerase chain reaction (PCR) in 29 KO inbred mouse strains via short tandem sequence repeat (STR) scanning and direct sequence cloning to justify microsatellite polymorphisms. The C57BL/6J and 129 mouse strains, from which these 29 KO mice were derived, were chosen as the background controls. The results indicated that 10 out of 42 (23.8%) loci showed CMP in some of these mouse strains. Except for the trinucleotide repeat locus of D3Mit22, which had microsatellite CMP in strain number 9, the core sequences of the remaining 41 loci were dinucleotide repeats, and 9 out of 41 (21.95%) showed CMPs among detected mouse strains. However, 11 out of 29 (37.9%) KO mice strains were recognized as having CMPs. The popular dinucleotide motifs in CMP were (TG)(n) (50%, 2/4), followed by (GT)(n) (27.27%, 3/11) and (CA)(n) (23.08%, 3/13). The microsatellite CMP in (CT)(n) and (AG)(n) repeats were 20% (1/5). According to cloning sequencing results, 6 KO mouse strains showed insertions of nucleotides whereas 1 showed a deletion. Furthermore, 2 loci (D13Mit3 and D14Mit102) revealed CMP in 2 strains, and mouse strain number 9 showed CMPs in two loci (D3Mit22 and D13Mit3) simultaneously. Collectively, these results indicated that microsatellite polymorphisms were present in the examined inbred KO mice.     

The AXB and BXA set of recombinant inbred mouse strains

The recombinant inbred (RI) set of strains, AXB and BXA, derived from C57BL/6J and A/J, originally constructed and maintained at the University of California/San Diego, have been imported into The Jackson Laboratory and are now in the 29th to 59th generation of brother-sister matings. Genetic quality control testing with 45 proviral and 11 biochemical markers previously typed in this RI set indicated that five strains had been genetically contaminated sometime in the past, so these strains have been discarded. The correct and complete strain distribution patterns for 56 genetic markers are reported for the remaining RI strain set, which consists of 31 living strains and 8 extinct strains for which DNA is available. Two additional strains, AXB 12 and BXA 17, are living and may be added to the set pending further tests of genetic purity. The progenitors of this RI set differ in susceptibility to 27 infectious diseases as well as atherosclerosis, obesity, diabetes, cancer, cleft palate, and hydrocephalus. Thus, the AXB and BXA set of RI strains will be useful in the genetic analysis of several complex diseases.     

Parental genetic contributions in the AXB and BXA recombinant inbred mouse strains

Recombinant inbred (RI) strains are a valuable tool in mouse genetics to rapidly map the location of a new locus. Because RI strains have been typed for hundreds of genetic markers, the genotypes of individual strains within an RI set can be examined to identify specific strain(s) containing the desired region(s) of interest (e.g., one or more quantitative trait loci, QTLs) for subsequent phenotype testing. Specific RI strains might also be identified for use as progenitors in the construction of consomic (chromosome substitution strains or CSSs) or congenic lines or for use in the RI strain test (RIST). To quickly identify the genetic contributions of the parental A/J (A) and C57BL/6J (B) strains, we have generated chromosome maps for each commercially available AXB and BXA RI strain, in which the genetic loci are colorcoded to signify the parent of origin. To further assist in strain selection for further breeding schemes, the percentages of A and B parental contributions were calculated, based on the total number of typed markers in the database for each strain. With these data, one can rapidly select the RI strain(s) carrying the desired donor and recipient strain region(s). Because points of recombination are known, starting with RI mice to generate CSSs or congenic lines immediately reduces genomewide screening to those donor-strain regions not already homozygous in the recipient strain. Two examples are presented to demonstrate potential uses of the generated chromosome maps: to select RI strains to construct congenic lines and to perform an RIST forAliq1, a QTL linked to ozone-induced acute lung injury survival.     

The LEXF: A new set of rat recombinant inbred strains between LE/Stm and F344

A new set of rat RI strains consisting of 11 independent strains and 13 of their substrains was established by inbreeding F₂ rats between F344/DuCrj and LE/Stm. The strain distribution pattern was examined for 66 microsatellite loci, 8 biochemical genetic markers, 2 histocompatibility loci, and 2 coat color genes. A rat salivary protein gene Spe1 was newly mapped on Chr 1. Received: 13 August 1996 / Accepted: 23 December 1996     

Effects of repeated maternal separation on prepulse inhibition of startle across inbred mouse strains

A growing body of research implicates genetic factors and childhood trauma in the etiology of neuropsychiatric diseases such as schizophrenia. However, there remains little understanding of how genetic variation influences early life stress to affect later disease susceptibility. Studies in rats have shown that postnatal maternal separation (MS) results in later deficits in prepulse inhibition of the acoustic startle response (PPI), an impairment in sensorimotor gating found in schizophrenic patients. In the present study, genetic differences in the effects of repeated MS on PPI were examined in eight inbred strains of mice (129S1/SvImJ, 129P3/J, A/J, BALB/cJ, BALB/cByJ C57BL/6J, DBA/2J and FVB/NJ). Mice were assigned to either MS (180 min/day on postnatal days P0-P13), 'handling' (15 min/day, P0-P13) or facility-reared conditions and tested for PPI at 12 weeks of age. Results demonstrated major strain differences in the production of viable offspring irrespective of MS, leading to the exclusion of 129P3/J, A/J and BALB/cJ from the study. Pups from the five remaining strains exhibited marked differences in the acoustic startle response and PPI, confirming previous strain comparisons. However, MS produced no significant effects on PPI in any of the strains tested. A second form of postnatal stress (repeated footshock) also failed to alter PPI in the one strain studied, C57BL/6J. Present results demonstrate that the form of MS studied herein does not provide a robust model of early life stress effects on PPI in the mouse strains tested. The development and validation of a reliable mouse model of early life stress remains an important research goal.     

Genetics of susceptibility to 6-aminonicotinamide-induced cleft palate in the mouse: studies in congenic and recombinant inbred strains

In a search for genetic differences in susceptibility to cleft palate, congenic and recombinant inbred strains of mice were treated with 6-aminonicotinamide or control injections. Of six loci tested, only the chromosome segment marked by N-acetyl transferase was found to affect susceptibility to 6-aminonicotinamide-induced cleft palate. This chromosome segment is known to affect glucocorticoid-induced cleft palate and phenytoin-induced cleft lip with or without cleft palate in these strains of mice.