Several investigators have reported genetic differences between sublines of highly inbred strains of mice. Bailey at this workshop has given considerable attention to the theoretical aspects of subline differentiation as these aspects may apply to the pedigrees of various strains and their sublines ( 1). Our particular study has for the most part concerned biochemical variants which are known to be polymorphic in feral populations and among strains which are relatively unrelated.
The data below have been obtained over about a 10-year period on strains which have been available at The Jackson Laboratory. In all cases, at least two individuals were examined from each strain to establish the allelotype of each locus. The loci chosen for study were dipeptidase-1 ( Dip-1), three esterases ( Es-1, Es-2, and Es-3), autosomal glucose-6-phosphate dehydrogenase ( Gpd-1), glucose phosphate isomerase-1 ( Gpi-1), hemoglobin β chain ( Hbb), isocitrate dehydrogenase-1 ( Id-1), lactate dehydrogenase regulator ( Ldr), supernatant malic enzyme ( Mod-1), two phosphoglucomutatases ( Pgm-1 and Pgm-2) and retinal degeneration ( rd). The last locus, which affects the retina of the eye, was chosen because the mutant allele is so frequently found among laboratory strains of mice, although it is probably not polymorphic in feral populations. in all analyses involving gel electrophoresis, controls with alternative alleles were present on each gel for purposes of comparison and ascertainment. References concerning electrophoretic procedures can be found in earlier studies ( 2). The strains listed for study have been described in various publications of Staats ( 3, 4).
The data are displayed in three tables. Table 1 comprises strains and sublines of the C57 groups whose relationships are shown in Figure 1. The relationships of the major groups are shown in the charts of Bailey ( 1). The several B-lines shown directly under C57BL/6J were derived from C57BL/6J and inbred by brother-sister mating. The RX-GE was a closed colony of mice derived from C57BL/10Gn and exposed to high doses of X-irradiation ( 5). Table 2 shows similar data for the DBA, BALB/c, A and related strains. Their relationships can be found in the pedigree chart of Staats ( 6). For our purposes, it is sufficient to point out that the A lines are related to BALB/c and that the C3H and CBA lines were derived from a cross between early progenitors of the DBA and BALB/c lines. Table 3 shows similar data for four unrelated groups. Their lack of relationship is based more on our lack of knowledge of their origins in feral populations. The S-lines were all brother-sister bred and were recent offshoots of SJL/J.
In all strains studied there was no evidence of within-strain genetic heterogeneity for those loci studied. This finding has been confirmed repeatedly by us with other enzyme polymorphisms. The lack of genetic heterogeneity within strains is not surprising probably to those attending this workshop, but it has been cause for surprise to investigators of other organisms, particularly those who study Drosophila. The constancy of genotype between distantly related sublines also supports the concept of genetic homozygosity of the strains studied.
The usefulness of each of the 13 loci studied depends on their respective degree of polymorphism in feral populations and among commonly used laboratory strains. For instance, a locus which displays a variant only rarely in feral populations or only in one or two laboratory strains would be less useful for this analysis than a locus where two alleles were in approximately equal frequencies among feral mice and among inbred strains. This reservation would pertain to analyses of both residual heterozygosity and mutation using this approach. Thus, loci Pgm-2, Ldr, Es-1, and Es-2 are less useful in differentiating strains than the other nine loci ( 2).
The data in Table 1 and Table 3 support the concept of consistency of sublines even as distantly separated as 200 generations. The genetic similarities between the major groups support the reality of their relationships as given by their purported pedigrees and further support the concept of genetic homogeneity and stability. These data confirm the close relationship of C57BL/Ks with other C57BL lines. This is in contrast to the data on histocompatibility differences shown by Graff ( 7) and leaves unresolved the question of origin or genetic contamination of this strain.
Table 2 provokes discussion. CBA/J differs from its related CBA/Ca lines at two loci, Pgm-1 and rd. And yet, CBA/J is identical with the C3H lines at these two loci as well as all others studied. Dr. Robert S. Bader who studied dentition patterns in these strains in the 1960s said that without doubt CBA/J and CBA/CaJ had similar dentition patterns suggesting their close relationship, but he did not mention his comparison of these patterns with C3H (Bader, personal communication). Green and Kaufer ( 8) found histocompatibility and other differences between CBA and CBA/Ca lines and stated that it would be advisable to regard the CBA strain as composed of at least two major sublines which should not be regarded as the same strain. Our data support their findings and recommendation. It is difficult to believe that residual heterozygosity would account for the differences at these two loci and at histocompatibility loci and for major differences in radiations sensitivity (cited in ref. 8). We interpret these data as supporting the hypothesis that the CBA line was at some time during its propagation in the United States since 1933 (the time of separation of the lines) inadvertently crossed with C3H with which it has often been housed, and with which it is outwardly identical.
Table 2 also reveals that DBA/1 and DBA/2 lines differ at the Gpd-1 locus. This is not surprising since these lines were not derived from a common inbred pair. We would interpret this difference as due to residual heterozygosity.
Sublines are for the most experimental purposes a nuisance because repeatability of experimental results depends in part on repeatability of the genotype of the experimental animal. So there are continuing attempts to minimize the number of sublines and the distances of relationships of experimental animals of a single inbred strain. With this paper, I wish to point out that there are good reasons for propagating at least some distantly related sublines. First, the analysis of subline differences to determine what heterogeneity persists or is evolving between lines is important to our concept of appropriate experimental animals to use. We need more information. If differences between sublines do appear, one can sometimes, especially with more than two branching sublines, determine from the pattern of alleles whether the variation is due to mutation or to residual heterozygosity. Thus, it is important to compare our observations of subline differentiation with Bailey's ( 1) theoretical expectations.
Second, one can use distantly related sublines for estimations of an upper limit of mutation rates under the assumption that most differences will be due to mutation and not residual heterozygosity. The denominator for estimating the rate for any locus can be based on a function of the number of individuals in each pedigree recognizing that new mutations, if neutral, will be fixed approximately one fourth of the time in a pedigree of sib-matings.
And third, if mutations do appear between closely related sublines, one can use the differing stocks in much the same way that congenic stocks have been used. Their utility is especially valuable in understanding the effects of a single locus without the problem of additional background genetic heterogeneity.
1. Bailey, D.W. (1978). In Origins of Inbred Mice (H.C. Morse III, ed.). Academic Press, New York.
2. Roderick, T.H., Ruddle, F.H., Chapman, V.M., and Shows, T.B. (1971). Biochem. Genet. 5: 457.
See also PubMed.
3. Staats, J. (1976). Cancer Res. 36: 4333.
See also MGI.
4. Staats, J. (1977). Inbred Strains of Mice No. 10. (This publication is issued as a companion to Mouse News Letter and has appeared
revised every other year since 1961. The earlier issues are indispensable for constructing pedigrees of inbred strains of mice).
See also MGI.
5. Green, E.L., and Les, E.P. (1964). Genetics 50: 497.
See also PubMed.
6. Staats, J. (1966). In Biology of the Laboratory Mouse (E.L. Green, ed.), p. 1. McGraw-Hill, new York.
7. Graff, R.J., (1970). Transplant. Proc. 2: 15.
See also MGI.
8. Green, M.C., and Kaufer, K.A. (1965). Transplantation 3: 766.
See also MGI.