Pathology of Genetically Engineered and Other Mutant Mice. Группа авторов

distributed evenly across all chromosomes, a pattern emerges in which markers specific to AKR/J and not BALB/cByJ skew to increased homozygosity in the region of the genome specific to the mutation. Usually this works relatively quickly to identify the region in which the mutated gene is located. However, in this particular case, less than 15% of the F2 mice were affected (Table 3.7) due to modifier genes in BALB/cByJ. Similar crosses with other strains resulted in the expected 25% distribution. This illustrates two points, the deviation from Mendelian expectation due to modifiers and the value of doing mapping crosses with different inbred strains, which in this instance identified different genetic intervals that together reduced the interval and the number of candidate genes (Figure 3.6) [9, 10].

Figure 3.5 Setting up a mapping cross using F1 and F2 hybrids.

      Backcrossing an F1 hybrid to one of the parents, instead of sibling intercrossing, is the initial process in the creation of a congenic, consomic, or conplastic strain, and such an N2 population is a useful tool for mapping dominant mutations. Some mutations or transgenes are adequately deleterious to overall health that they cannot be maintained on an inbred background and instead require hybrid vigor. Such mutants are maintained by breeding to an F1 hybrid at each generation or through outcross–intercross to an F1 hybrid. One complicated example of this is B6EiC3Sn a/A‐Ts(17¹⁶)65Dn/J [11]. This chromosomal aberration was induced in DBA/2J inbred mice, but is detrimental to health and causes male sterility so in order to successfully maintain a colony a female carrier is bred to a (C57BL/6JEiJ x C3H/HeSnJ)F1/J male at each generation. There is no punctuation in the genetic background of the strain name, B6EiC3Sn, and the presence of the DBA/2J contribution is omitted from the nomenclature for simplicity and must be found in sources outside of the strain nomenclature.

Table 3.7 Breeding data used to identify the gene responsible for the hair interior defect phenotype.

AKR/J‐hid* homozygote crossed with	Total progeny	Normal mice	Affected mice	Affected (%)	X 2	p‐Value
BALB/cByJ	260	222	38	14.62	5.92	0.025–0.010
C57BL/6J	430	330	100	23.26	0.220	0.500–0.750
CAST/EiJ	339	255	84	24.78	0.003	>0.900
C3H/HeJ	248	191	57	22.98	0.170	0.500–0.750
FVB/NJ	272	205	67	24.63	0.006	>0.900
Total F2 mice	1549	1203	346

      ^* AKR/J‐hid was the original designation until the gene responsible was identified. Because the gene was identified with a different phenotype earlier, the allele is officially Soat1^ald not Soat1^hid.

Figure 3.6 Crossing AKR/J mice with more than one inbred strain to map the mutant gene locus. By using the shortest interval between different crosses, the list of candidate genes could be reduced. Sequencing then only had to be done on a few genes to identify the mutant gene responsible for the hair interior defect phenotype.

      Sources: Based on Giehl KA et al. [9].

Some mutations are maintained on a background in which the genetic contributions of two inbred strains are segregating. The retarded hair growth (in ornithine aminotransferase, Oat^rhg) mutation arose spontaneously in AKR/J and was later outcrossed to C57BL/6JEi and subsequently maintained by sibling intercrossing [12, 13]. Thus, the genetic background consists of contributions from both AKR/J and C57BL/6JEi segregating at each generation. A semicolon between the strain abbreviations is used to represent this situation, B6Ei;AKR‐Oat^rhg/J. If more than two inbred strains contribute to the genetic background of a strain, then it is simply called a stock, such as STOCK Noto^tc/J (notochord homeobox gene, truncate mutation). The truncate mutation arose spontaneously in the F4 generation of a cross between a C3H female and a Swiss outbred male. It was subsequently bred to BALB/cHu and again to C3H before being inbred for more than 80 generations [14]. The STOCK is in capital letters designating inbred, and there is no hyphen between STOCK and the allele symbol. The strain could have been assigned a unique inbred strain name, such as TC/J, but was instead designated as a STOCK. When stock is in lower case, it indicates that the segregating population has not reached F20.

Recombinant Inbred (RI) and Recombinant Congenic Mice

One variation of inbred mice is the creation of recombinant inbred (RI) mice. In this case, two different inbred strains are crossed to produce F1 progeny. As with creating inbred strains, breeding pairs of these F1 progeny are crossed to create an F2 population and then one pair is selected for breeding in this and each subsequent generation until at 20 consecutive generations of breeding (F20) from a single brother–sister pair a new inbred strain is created, which has approximately 50% of its DNA derived from each of the original parental inbred strains. The generation of many separate recombinant inbred strains from the same parental inbred strains creates a panel of recombinant inbred stains, each of which has an array of unique DNA recombinations and therefore unique subset of parental DNA segments. Recombinant inbred strains are named with a pair of 1‐ or 2‐letter strain abbreviation codes representing the original parental inbred strains (Figure 3.3) with the female founder strain first, followed by an X for the cross, then the male founder strain abbreviation followed by a slash, a number to indicate the specific line and then the laboratory code for who made the strain (Table 3.8). The higher the number of strains in a recombinant inbred panel, the potentially more valuable the collection due to the increased number of recombination sites