Applications of congenic strains in the mouse

The sequencing of the human and the mouse genomes has shown that the chromosomes of these two species contain approximately 30,000 genes. The biological systems that can be studied in an individual or in a tissue result from complex interactions within this multitude of genes. Before describing these interactions, it is necessary to understand the function of each gene. In the mouse, congenic strains are developed to introduce a chromosomal segment in a given inbred genetic background. One can then compare the biological effects of different alleles at the same locus in the same genetic background or the effect of a given allele in different genetic backgrounds. One can also introduce into different congenic strains with the same genetic background genes which control a complex genetic trait, then combine these genes by appropriate crosses to study their interactions. Although the chromosomal segment transferred into a congenic strain usually contains up to several hundreds of genes, molecular markers can be used to reduce this number as well as the number of crosses required for the development of congenic strains.     

Possible Derivation of the Laboratory Mouse Genome from Multiple Wild MUS Species

Laboratory strains of mice are thought to be derived from wild populations of Mus domesticus. Many instances of non-domesticus genetic information fixed in these strains have been described, however, and the amount of strain-to-strain genetic variation exceeds that found in wild domesticus populations. In order to estimate the extent of the non-domesticus contribution to laboratory mouse genomes, and to determine whether it could account for observed variation, we have used computer simulations to investigate the properties of genetically marked chromosomal segments and the distribution of residual allogenicity at various times during inbreeding. A locus or chromosomal segment is allogenic if it is unfixed within a lineage at a given time. The odds of fixation of a foreign chromosome segment are predicted to be an exponentially decreasing function of its length. The median segment length is predicted to be 17 centimorgans. Available data for markers of chromosomes 1, 9 and 12 in recombinant inbred strain sets conform to these predictions. Together, the results suggest that introgression of non-domesticus chromosomes and segregation of residual allogenicity are sufficient to account for the genetic diversity observed among inbred mouse strains and substrains.     

Novel insights into the genetic background of genetically modified mice

Unclear or misclassified genetic background of laboratory rodents or a lack of strain awareness causes a number of difficulties in performing or reproducing scientific experiments. Until now, genetic differentiation between strains and substrains of inbred mice has been a challenge. We have developed a screening method for analyzing inbred strains regarding their genetic background. It is based on 240 highly informative short tandem repeat (STR) markers covering the 19 autosomes as well as X and Y chromosomes. Combination of analysis results for presence of known C57BL/6 substrain-specific mutations together with autosomal STR markers and the Y-chromosomal STR-haplotype provides a comprehensive snapshot of the genetic background of mice. In this study, the genetic background of 72 mouse lines obtained from 18 scientific institutions in Germany and Austria was determined. By analyzing only 3 individuals per genetically modified line it was possible to detect mixed genetic backgrounds frequently. In several lines presence of a mispairing Y chromosome was detected. At least every second genetically modified line displayed a mixed genetic background which could lead to unexpected and non-reproducible results, irrespective of the investigated gene of interest.     

Simulation of the distribution of parental strains’ genomes in RC strains of mice

Recombinant Congenic strains (RC strains) were developed to facilitate mapping of genes influencing complex traits controlled by multiple genes. They were produced by inbreeding of the progeny derived from a second backcross from a common `donor' inbred strain to a common `background' inbred strain. Each RC strain contains a random subset of approximately 12.5% of genes from the donor strain and 87.5% of genes from the background strain. In this way the genetic control of a complex disease may be dissected into its individual components. We simulated the production of the RC strains to study to what extent they have to be characterized in order to obtain sufficient information about the distribution of the parental strains' genomes in these strains and to acquire insight into parameters influencing their effectiveness in mapping quantitative trait loci (QTLs). The donor strain genome in the RC strains is fragmented into many segments. Genetic characterization of these strains with one polymorphic marker per 3.3 centiMorgans (cM) is needed to detect 95% of the donor strain genome. The probability of a donor strain segment being located entirely in between two markers of background strain origin that are 3 cM apart (and hence escaping detection) is 0.003. Although the donor strain genome in the RC strains is split into many segments, the largest part still occurs in relatively long stretches that are mostly concentrated in fewer than 13 autosomes, the median being 9 autosomes. Thus, in mapping QTLs, the use of RC strains facilitates the detection of linkage. Received: 20 December 1996 / Accepted: 23 July 1997     

Sex, strain, and species differences affect recombination across an evolutionarily conserved segment of mouse chromosome 16

A region of substantial genetic homology exists between human chromosome 21 (HSA21) and mouse chromosome 16 (MMU16). Analysis of 520 backcross animals has been used to establish gene order in the homologous segment. D21S16h and Mx are shown to represent the known proximal and distal limits of homology between the chromosomes, while Gap43, whose human homolog is on HSA3, is the next proximal marker on MMU16 that has been mapped in the human genome. Recombination frequencies (RFs) in four intervals defined by five loci in the HSA21-homologous region of MMU16 were analyzed in up to 895 progeny of eight different backcrosses. Two of the eight crosses were made with F1 males and six with F1 females. The average RF of 0.249 in 265 backcross progeny of F1 males was significantly higher than the 0.106 average recombination in 320 progeny of F1 females in the interval from D21S16h to Ets-2. This is in contrast to HSA21, which shows higher RFs in female meiosis in the corresponding region. Considerable variation in RF was observed between crosses involving different strains, both in absolute and in relative sizes of the intervals measured. The highest RFs occurred in a cross between the laboratory strain C57BL/6 and MOLD/Rk, an inbred strain derived from Mus musculus molossinus. RFs on this cross were nearly fivefold higher than those reported previously for an interspecific cross between C57BL/6 and Mus spretus.     

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.     

A novel strategy for genetic dissection of complex traits: the population of specific chromosome substitution strains from laboratory and wild mice

The mouse is an irreplaceable model for understanding the precise genetic mechanisms of mammalian physiological pathways. Thousands of quantitative trait loci (QTLs) have been mapped onto the mouse genome during the last two decades. However, only a few genes’ underlying complex traits have been successfully identified, and rapid fine mapping of QTL genes still remains a challenge for mouse geneticists. Currently, the Collaborative Cross (CC) has proceeded to the goal of establishing more than 1,000 recombinant inbred strains for the sub-centimorgan mapping resolution of QTL loci. In this article, a novel complementary strategy, designated as population of specific chromosome substitution strains or PSCSS, is proposed for rapid fine mapping of QTLs on the substituted chromosome. One specific chromosome (Chr 1) of recipient mouse strain C57BL/6 J has been substituted by homologous counterparts from five different inbred strains (C3H/He, FVB/N, AKR, NOD/LtJ, NZW/LacJ), an outbred line Kunmin mouse in China, and 23 wild mice captured in different localities. The primary genetic studies on the structure of these wild donor chromosomes (Chr 1) show that these donor chromosomes harbor extensive genetic polymorphisms, with a high density of SNP distribution, abundant variations of STR alleles, and a high level of historical recombination accumulation. These specific chromosome substitution strains eventually form a special population that has the identical genetic background of the recipient strain and differs only in the donor chromosomes. With simple association studies, known QTLs on the donor chromosome can be rapidly mapped in high resolution without requirement of further crosses. This approach, taking advantage of the extensive genetic polymorphisms of wild resources and chromosome substitution strategy, brings a new outlook for genetic dissection of complex traits.     

Systematic Identification of Balanced Transposition Polymorphisms in Saccharomyces cerevisiae

High-throughput techniques for detecting DNA polymorphisms generally do not identify changes in which the genomic position of a sequence, but not its copy number, varies among individuals. To explore such balanced structural polymorphisms, we used array-based Comparative Genomic Hybridization (aCGH) to conduct a genome-wide screen for single-copy genomic segments that occupy different genomic positions in the standard laboratory strain of Saccharomyces cerevisiae (S90) and a polymorphic wild isolate (Y101) through analysis of six tetrads from a cross of these two strains. Paired-end high-throughput sequencing of Y101 validated four of the predicted rearrangements. The transposed segments contained one to four annotated genes each, yet crosses between S90 and Y101 yielded mostly viable tetrads. The longest segment comprised 13.5 kb near the telomere of chromosome XV in the S288C reference strain and Southern blotting confirmed its predicted location on chromosome IX in Y101. Interestingly, inter-locus crossover events between copies of this segment occurred at a detectable rate. The presence of low-copy repetitive sequences at the junctions of this segment suggests that it may have arisen through ectopic recombination. Our methodology and findings provide a starting point for exploring the origins, phenotypic consequences, and evolutionary fate of this largely unexplored form of genomic polymorphism.     

Hippocampal commissure defects in crosses of four inbred mouse strains with absent corpus callosum

It is known that four common inbred mouse strains show defects of the forebrain commissures. The BALB/cJ strain has a low frequency of abnormally small corpus callosum, whereas the 129 strains have many animals with deficient corpus callosum. The I/LnJ and BTBR T+ tf/J strains never have a corpus callosum, whereas half of I/LnJ and almost all BTBR show severely reduced size of the hippocampal commissure. Certain F1 hybrid crosses among these strains are known to be less severely abnormal than the inbred parents, suggesting that the parent strains have different genetic causes of commissure defects. In this study, all hybrid crosses among the four strains were investigated. The BTBR × I/Ln hybrid expressed almost no defects of the hippocampal commissure, unlike its inbred parent strains. Numerous three‐way crosses among the four strains yielded many mice with no corpus callosum and severely reduced hippocampal commissure, which shows that the phenotypic defect can result from several different combinations of genetic alleles. The F2 and F3 hybrid crosses of BTBR and I/LnJ had almost 100% absence of the corpus callosum but about 50% frequency of deficient hippocampal commissure. The four‐way hybrid cross among all four abnormal strains involved highly fertile parents and yielded a very wide phenotypic range of defects from almost no hippocampal commissure to totally normal forebrain commissures. The F2 and F3 crosses as well as the four‐way cross provide excellent material for studies of genetic linkage and behavioral consequences of commissure defects.     

Mouse phenome research: implications of genetic background

Now that sequencing of the mouse genome has been completed, the function of each gene remains to be elucidated through phenotypic analysis. The "genetic background" (in which each gene functions) is defined as the genotype of all other related genes that may interact with the gene of interest, and therefore potentially influences the specific phenotype. To understand the nature and importance of genetic background on phenotypic expression of specific genes, it is necessary to know the origin and evolutionary history of the laboratory mouse genome. Molecular analysis has indicated that the fancy mice of Japan and Europe contributed significantly to the origin of today's laboratory mice. The genetic background of present-day laboratory mice varies by mouse strain, but is mainly derived from the European domesticus subspecies group and to a lesser degree from Asian mice, probably Japanese fancy mice, which belong to the musculus subspecies group. Inbred laboratory mouse strains are genetically uniform due to extensive inbreeding, and they have greatly contributed to the genetic analysis of many Mendelian traits. Meanwhile, for a variety of practical reasons, many transgenic and targeted mutant mice have been created in mice of mixed genetic backgrounds to elucidate the function of the genes, although efforts have been made to create inbred transgenic mice and targeted mutant mice with coisogenic embryonic stem cell lines. Inbred mouse strains have provided uniform genetic background for accurate evaluation of specific genes phenotypes, thus eliminating the phenotypic variations caused by mixed genetic backgrounds. However, the process of inbreeding and selection of various inbred strain characteristics has resulted in inadvertent selection of other undesirable genetic characteristics and mutations that may influence the genotype and preclude effective phenotypic analysis. Because many of the common inbred mouse stains have been established from relatively small gene pools, common inbred strains have limitations in their genetic polymorphisms and phenotypic variations. Wild-derived mouse strains can complement deficiencies of common inbred mouse strains, providing novel allelic variants and phenotypes. Although wild-derived strains are not as tame as the common laboratory strains, their genetic characteristics are attractive for the future study of gene function.     

Mixed Effects Models for Quantitative Trait Loci Mapping With Inbred Strains

Fixed effects models have dominated the statistical analysis of genetic crosses between inbred strains. In spite of their popularity, the traditional models ignore polygenic background and must be tailored to each specific cross. We reexamine the role of random effect models in gene mapping with inbred strains. The biggest difficulty in implementing random effect models is the lack of a coherent way of calculating trait covariances between relatives. The standard model for outbred populations is based on premises of genetic equilibrium that simply do not apply to crosses between inbred strains since every animal in a strain is genetically identical and completely homozygous. We fill this theoretical gap by introducing novel combinatorial entities called strain coefficients. With an appropriate theory, it is possible to reformulate QTL mapping and QTL association analysis as an application of mixed models involving both fixed and random effects. After developing this theory, our first example compares the mixed effects model to a standard fixed effects model using simulated advanced intercross line (AIL) data. Our second example deals with hormone data. Here multivariate traits and parameter identifiability questions arise. Our final example involves random mating among eight strains and vividly demonstrates the versatility of our models.     

Ribosomal DNA inheritance and recombination in Neurospora crassa

Summary The genetic segregation of ribosomal DNA (rDNA) in Neurospora crassa was analyzed by exploiting restriction fragment length polymorphisms in the nontranscribed spacer (NTS) sequences of nine laboratory wild-type strains and wild-collected strains. In an analysis of random spore progeny from seven crosses, and of ordered tetrads from two of those crosses the rDNA was shown to be inherited in a simple, stable Mendelian fashion, exhibiting an approximately 1:1 ratio of the two parental rDNA types. No meiotic recombinants were detected among the progeny, indicating that non-sister-chromatid crossing over is highly suppressed in the rDNA region. The basis for this suppression of meiotic recombination is not known.     

Use of a novel outbred by inbred F1 cross to detect genetic markers for growth

A unique outbred by inbred F1 resource population was established. The population structure facilitated the unique opportunity of examining gene by genetic background interaction through crossing two modern broiler sires with dams from two unrelated inbred lines, with no selection for growth rate, to produce about 600 F1 chicks. Pools of DNA were generated from the phenotypic extremes (20% high and low) for 8-week body weight for each of the four combinations of sire and dam line. For one sire family, pools were also separately generated for each sex. The pools were genoyped with 25 informative (segregating) microsatellites. This unique F1 cross between outbred and inbred populations allowed use of the inbred alleles as an 'internal control' for polymerase chain reaction amplification quality in DNA pools. Ten microsatellites showed marked differences (P < 0.05) in allele frequencies between high and low pools, suggesting an association between marker and quantitative trait loci (QTL). These differences were verified using selective genotyping. For many markers, differences in allele frequencies between the high and the low pools, or marker effect, varied between the two dam lines and the two sexes, suggesting an interaction between some genes and the genetic background as represented by different dam lines or sexes. The suggestive marker-QTL associations identified in this F1 population demonstrate the efficiency of this population design while different QTL effects in different genetic line crosses and sexes highlight the importance of gene by genetic background interaction in QTL detection.     

Genetic variation in dosage effects in maize aneuploids.

In maize (Zea mays L.), the consequences of aneuploidy have been well documented, however, genetic variation in the responses to aneuploidy has not been examined. Using simple B-A translocation stocks to generate a dosage series involving segments from 14 chromosome arms, we tested for the presence of genetic variation for dosage responses in maize by examining reciprocal and maternal genotype effects on the dosage responses. Reciprocal effects examined whether there were differences between two distinctly different inbred backgrounds, Mo17Ht and B73Ht, in how they responded to loss or gain of a B73Ht segment in the Mo17Ht x B73Ht (TB) F1 cross versus a Mo17Ht segment in the B73Ht x Mo17Ht (TB) F1 cross. Maternal genotype effects questioned whether different inbred backgrounds, Sc41R, T8, and either Mo17Ht or B73Ht (depending on the male), when used as females responded differently to the loss or gain of a chromosome arm segment from the same male (either B73Ht TB or Mo17Ht TB) in an F1 cross. Numerous examples of reciprocal and maternal genetic effects were identified in this study. Most of the genetic effects were due to differences in magnitude of response rather than direction; however, tassel-branch number involving the 5S chromosome segment in the B73Ht male background and the 7L chromosome segment in the Mo17Ht male background showed a trend toward the maternal genotype effects being due to differences in the direction of the response. Key words : quantitative traits, corn, B-A translocations, dosage analysis.     

Mating preferences of F2 segregants of crosses between MHC-congenic mouse strains

Male and female F₂ homozygotes from crosses between MHC-congenic inbred mouse strains were tested for MHC-associated mating preference. In three instances, of the four genotypic combinations so tested, marked MHC-associated mating preference was observed. This result greatly reduces the possibility that the observed mating preferences of MHC-congenic inbred strains can be explained wholly in terms of non-MHC genetic drift, or of residual non-MHC genetic disparity, or of fortuitous acquired strain characteristics unrelated to MHC. In two of the four combinations investigated, the MHC-related mating bias of F₂ segregants was similar to that of the genotypically similar inbred parent strains. In a third combination, F² segregants did not show the mating bias exhibited by the corresponding parent strains. In a fourth combination, F₂ segregants displayed an MHC-related mating bias that was evident in the corresponding parental inbred strains only when the colonies of the parent strains had been maintained in isolation from other strains. While the exhibition of mating preference by mice of the same genotypes may differ according to circumstances, as indicated, in no instance was preference reversed. Mating preference in a given combination of MHC genotypes, whenever it was observed, always favored the same MHC haplotype of the two alternative haplotypes represented. It appears that the familial MHC genotypes of mice and the environment in which the colonies are maintained influence their MHC-related mating preference, but it has yet to be decided whether these factors operate by determining exposure to particular MHC haplotypes.Abbreviations used in this paper are as follows B6 C57BL/6 - B10 C57BL/10 - BALE BALB/c - BALB.B BALB.B10 - INB inbred - MHC major histocompatibility complex See also Figure 1     

Gene conversion in nonsense suppressors of Schizosaccharomyces pombe. I. The influence of the genetic background and of three mutant genes (rad2, mut1 and mut2) on the frequency of the post-meiotic segregation.

Summary The frequency of gene conversion was assessed at the anticodon site of the sup3 gene of Schizosaccharomyces pombe. In order to detect a possible influence of the genetic background on the relative frequency of post-meiotic segregations amongst conversions, three similar crosses were analyzed which differed as follows: In cross I the two parents were derived by spontaneous mutation from one and the same strain. The heterogeneity of the genetic background between these two parent strains is assumed to be at a minimum level. In cross II the crossing partners were strains of the Bernese stock collection and differ probably in their genetic background to some extent. Last, in cross III, one of the parent strains was ten times repeatedly mutagenized with nitrosoguanidine in order to introduce cryptic mutations in the genome. A maximum degree of background heterogeneity between parents is expected for this cross. Neither the total conversion frequency nor the frequency of post-meiotic segregations amongst conversions were found to be significantly different in these three crosses.In addition, no effect of the radiation-sensitive mutation, rad2-44, and of two mutator mutations, mut1-4 and mut2-9, on the conversion pattern could be detected.     

Major Effects on Teratogen-Induced Facial Clefting in Mice Determined by a Single Genetic Region

A major correlation has been found between the incidence of glucocorticoid-induced cleft palate and the chromosome 8 segment identified by N-acetyl transferase in mice. The resistant strain became fully susceptible while the susceptible strain became resistant when this chromosomal region, representing less than 0.7% of the genome, was transferred from one strain to the other by the construction of congenic strains. 6-Aminonicotinamide-induced cleft palate and phenytoin-induced cleft lip with or without cleft palate are also influenced by this genetic region but not as strongly. In both cases the susceptible strain became quite resistant to the teratogen-induced clefting when the N-acetyl transferase region of chromosome 8 was transferred. However, this chromosomal region does not make the resistant strain susceptible to these two teratogens.     

Magnaporthe Grisea Genes for Pathogenicity and Virulence Identified through a Series of Backcrosses

We have identified genes for pathogenicity toward rice (Oryza sativa) and genes for virulence toward specific rice cultivars in the plant pathogenic fungus Magnaporthe grisea. A genetic cross was conducted between the weeping lovegrass (Eragrostis curvula) pathogen 4091-5-8, a highly fertile, hermaphroditic laboratory strain, and the rice pathogen O-135, a poorly fertile, female-sterile field isolate that infects weeping lovegrass as well as rice. A six-generation backcrossing scheme was then undertaken with the rice pathogen as the recurrent parent. One goal of these crosses was to generate rice pathogenic progeny with the high fertility characteristic of strain 4091-5-8, which would permit rigorous genetic analysis of rice pathogens. Therefore, progeny strains to be used as parents for backcross generations were chosen only on the basis of fertility. The ratios of pathogenic to nonpathogenic (and virulent to avirulent) progeny through the backcross generations suggested that the starting parent strains differ in two types of genes that control the ability to infect rice. First, they differ by polygenic factors that determine the extent of lesion development achieved by those progeny that infect rice. These genes do not appear to play a role in infection of weeping lovegrass because both parents and all progeny infect weeping lovegrass. Second, the parents differ by simple Mendelian determinants, ``avirulence genes,' that govern virulence toward specific rice cultivars in all-or-none fashion. Several crosses confirm the segregation of three unlinked avirulence genes, Avr1-CO39, Avr1-M201 and Avr1-YAMO, alleles of which determine avirulence on rice cultivars CO39, M201, and Yashiro-mochi, respectively. Interestingly, avirulence alleles of Avr1-CO39, Avr1-M201 and Avr1-YAMO were inherited from the parent strain 4091-5-8, which is a nonpathogen of rice. Middle repetitive DNA sequences (``MGR sequences'), present in approximately 40-50 copies in the genome of the rice pathogen parent, and in very low copy number in the genome of the nonpathogen of rice, were used as physical markers to monitor restoration of the rice pathogen genetic background during introgression of fertility. The introgression of highest levels of fertility into the most successful rice pathogen progeny was incomplete by the sixth generation, perhaps a consequence of genetic linkage between genes for fertility and genes for rice pathogenicity. One chromosomal DNA segment with MGR sequence homology appeared to be linked to the gene Avr1-CO39. Finally, many of the crosses described in this paper exhibited a characteristic common to many crosses involving M. grisea rice pathogen field isolates. That is, pigment-defective mutants frequently appeared among the progeny.     

Mouse inbred strain sequence information and yin-yang crosses for quantitative trait locus fine mapping

The shared ancestry of mouse inbred strains, together with the availability of sequence and phenotype information, is a resource that can be used to map quantitative trait loci (QTL). The difficulty in using only sequence information lies in the fact that in most instances the allelic state of the QTL cannot be unambiguously determined in a given strain. To overcome this difficulty, the performance of multiple crosses between various inbred strains has been proposed. Here we suggest and evaluate a general approach, which consists of crossing the two strains used initially to map the QTL and any new strain. We have termed these crosses "yin-yang," because they are complementary in nature as shown by the fact that the QTL will necessarily segregate in only one of the crosses. We used the publicly available SNP database of chromosome 16 to evaluate the mapping resolution achievable through this approach. Although on average the improvement of mapping resolution using only four inbred strains was relatively small (i.e., reduction of the QTL-containing interval by half at most), we found a great degree of variability among different regions of chromosome 16 with regard to mapping resolution. This suggests that with a large number of strains in hand, selecting a small number of strains may provide a significant contribution to the fine mapping of QTL.     

The interaction of two groups of murine genes determines the persistence of Theiler''s virus in the central nervous system.

Theiler's murine encephalomyelitis virus is responsible for a chronic inflammatory demyelinating disease of the central nervous system of the mouse. The disease is associated with persistent viral infection of the spinal cord. Some strains of mice are susceptible to viral infection, and other strains are resistant. The effect of the genetic background of the host on viral persistence has not been thoroughly investigated. We studied the amount of viral RNA in the spinal cords of 17 inbred strains of mice and their F1 crosses with the SJL/J strain and observed a large degree of variability among strains. The pattern of viral persistence among mouse strains could be explained by the interaction of two loci. One locus is localized in the H-2D region of the major histocompatibility complex, whereas the other locus is outside this complex and is not linked to the Tcrb locus on chromosome 6.