B6ad

Figure 9.4 Construction of double congenic mouse strains.

is used to assess the effects of one genetic region on another as illustrated in Figure 9.5. Thus, differences between B6 and B6.A.D compared to those between B6 and B6.A will disclose the effect of a segment of chromosome 12 on the expression of the differential Nat allele on chromosome 8. Conversely, differences between B6 and B6.A.D compared to those between B6 and B6.D will disclose the effect of a segment on chromosome 8 on the expression of the differential Ahr receptor allele on chromosome 12.

Recombinant inbred strains together with congenic strains have been used with great success to investigate a number of qualitative and some quantitative genetically polymorphic traits. These systems are most effective for analysis of traits whose characteristics are determined primarily by a single gene. They can help determine whether more than one gene is involved, and the minimal number that is involved, but cannot identify these loci. Hence they are less effective in studying multifactorial traits.

Recombinant Congenic Strains

In more complex systems, additive and interactive effects obscure or disrupt the association of genotype and phenotype that is required to interpret observed SDPs of recombinant inbred lines. As a means of mapping and investigating the individuality of genes that play a role in multifactorial traits, recombinant congenic strains were developed. In this system, nonlinked genes are sep-

Strain B6

Strain B6.D

Chr 8 Chr 12

Strain B6.A

D allele difference on chromosome 12 J*

Chr 8 plus Chr 12 differences

Chr 8 Chr 12

Strain B6.A.D

D allele difference on chromosome 12

Natr vs Nats difference on chromosome 8

Natr vs Nats difference on chromosome 8

Figure 9.5 Quartets of double congenic mouse strains.

arated in different recombinant congenic strains and are then more accessible to study.

The procedure for producing recombinant congenic strains combines the approaches described above for the construction of recombinant inbred strains and congenic strains. By limited backcrossing of two selected strains and subsequent brother-sister mating, a set of 15-20 strains is produced. Each of the individual strains carries a small fraction of the genome of the donor strain on the genome of the background strain. The number of backcross generations used, unlike that for the production of strains congenic at a single locus, depends on the degree of genetic resolution called for to separate the genes of interest that are transferred. For example, two backcross generations yield an average of 12.5% of the donor genome transferred and 87.5% of the background genome retained, while three backcrosses result in the transfer of 6.25% of the donor genome. Consequently, genes from the donor that are unlinked are likely to be redistributed into different strains. A multigene trait that depends on differential alleles at multiple loci is thereby transformed into a set of single gene differences between the background strain and individual recombinant congenic strains.

Recombinant congenic strains provide a versatile experimental system for separating unlinked loci affecting a given trait, for establishing new linkage relationships for any trait with genes that can be assayed, and if appropriate crosses are made between recombinant congenic strains carrying different genes, they can provide a flexible system for evaluating gene interactions. When recombinant congenic strains are used in combination with mice created by gene targeting (see p. 278), they can be used to assess the influence of the rest of the genome on the expression and function of the targeted gene. In this way, recombinant congenic strains and strains created by gene targeting complement each other. The primary disadvantage of recombinant congenic strains is the same as that for recombinant inbred strains—they can be used to analyze only genes that possess differential alleles at loci of interest.

A considerable amount of information can be collected by polymerase chain reaction in a short time for SSLPs. The SDPs of SSLPs in a set of recombinant congenic strains can then be used to search the mouse genome for new genes involved in the control of polygenic traits. For example, the SDPs of SSLPs in a CcS/Dem set of recombinant congenic strains have been used to search for genes that are involved in the genetic control of colon cancer. The parental strains of the CcS/Dem set, BALB/cHea (background strain) and STS/A (donor strain), differ in the number of colon tumors induced by the carcinogen 1,2-dimethylhydrazine: BALB/c mice are relatively resistant and STS mice are highly sensitive to tumor induction by this agent. Among the set of 20 CcS/Dem recombinant congenic strains created from these parental lines, several were highly sensitive and several were resistant. Each CcS/Dem strain carries a unique subset of about 12.5% of the genes derived from the STS strain on the BALB/c background, and individual STS susceptibility genes are segregated into different recombinant con-genic strains. To map the susceptibility of gene(s) present in one of the highly susceptible strains, CcS-19, BALB/c x (BALB/c x CsS-19) Fi backcross mice were treated with 1,2-dimethylhydrazine. After 6 weeks, the number and size of colon tumors were recorded, and linkage of the susceptibility to the battery of SSLP markers for the STS allele A was tested. A new susceptibility gene (Scc-1) for colon tumors was mapped to chromosome 2 in the vicinity of the SSLP marker. CD44. Scc-1 differs from the oncogenes and tumor suppressor genes known to be involved in colon tumorigenesis. Because the colon tumors induced were significantly larger than those in BALB/c mice, and because the size of the tumors did not segregate with CD44, the investigators also concluded that the number and size of the tumors are controlled by different genes.

Chromosome Substitution Strains (CSSs)

CSSs were originally proposed as an approach to gene identification of complex traits by quantitative trait locus (QTL) analysis17 (see p. 282). A CSS is an inbred strain in which one chromosome has been substituted from a different inbred strain by repeated backcrossing. The CSS A.B-Chr(i) is thus defined as an inbred strain that is identical to strain A except that chromosome "i" has been substituted by the corresponding chromosome from strain B. In the mouse, a CSS panel consists of 21 strains, corresponding to the 19 autosomes and two sex chromosomes.

The strategy for constructing a CSS as described by Nadeau et al.17 is shown in Figure 9.6. As a pilot project to test the usefulness of CSSs to dissect genetic factors affecting complex traits, these investigators constructed a complete CSS panel

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