Pathology of Genetically Engineered and Other Mutant Mice. Группа авторов

across the panel.

Table 3.8 Recombinant inbred and recombinant congenic strain designation.

BXD31/TyJ	AcB51/Prgs
B = C57BL/6J	A = backcross to A/J
X = cross	c = recombinant congenic cross
D = DBA/2J	B = C57BL/6J
31 = RI line number	51 = recombinant congenic line number
Ty = Lab that created the line (Benjamin Taylor)	Prgs = Lab that created the line (Philippe Gros)
J = Subsequent breeder of the strain, The Jackson Laboratory

The Jackson Laboratory Repository distributes more than 125 BXD RI lines and there are many more in private laboratories. Many of these BXD RI lines have been sequenced and annotated. Using websites, such as GeneNetwork (www.genenetwork.org) [15], it is possible to use these mice for localizing, if not necessarily identifying, the gene(s) responsible for a disease, response to a treatment, or for any phenotype observed in one of the parental strains but not the other. For example, if the primary gene responsible for a disease in DBA/2J mice is known, such as a hypomorphic allele of Abcc6 that causes pseudoxanthoma elasticum, and the gene is wildtype in C57BL/6J, by first identifying all the BXD lines that are homozygous for the D2 mutant allele, one can then select only those mice, phenotype them, and rapidly identify narrow genetic intervals from which to generate a small list of candidate disease modifier genes [16] (Figure 3.7). Other RI lines exist for a variety of crosses that can be used in a similar manner.

      A variation on the RI line is the recombinant congenic strains that are created by crossing two inbred strains followed by one or several backcrosses to one of the parental strains. The resulting mice are then inbred without selection. Recombinant congenic strains are also named using 1‐ or 2‐letter strain abbreviation codes representing the parental strains, but a lower case c for congenic, not an X, separates them and the first strain listed is the host strain, that to which the backcrossing is done, and the strain symbol following the c is the donor strain, that contributing the least amount to the total genetic background. The AcB51/Pgrs recombinant congenic strain was generated by Dr. Philippe Gros by backcrossing (A/J x C57BL/6J)F1 to A/J for two consecutive generations then sibling inbreeding to generate a panel of recombinant consomics including this, line 51.

Collaborative Cross (CC) Mice

      Another variation of RI lines is the Collaborative Cross. This is a complex set of crosses using eight different inbred strains that represent a great deal of genetic diversity [17, 18]. The goal was to create over 1000 new inbred strains. Due to reproductive failure and low viability of many strains, the current goal is to create 200 strains. These mice present with great phenotypic variability, a reflection of the great genetic diversity. These can be used to screen for specific phenotypes [19, 20]. When a large enough cohort are examined, phenotyped, and genotyped using the Mouse Universal Genotyping Array (Single Nucleotide Polymorphism [SNP] Array), it is possible to identify candidate gene regions and even the gene(s) responsible for the phenotype [21].

      Collaborative Cross mice are designated by CC for Collaborative Cross, the line number, forward slash, and the lab that created it followed by the laboratory that currently maintains it. For example, CC001/UncJ (stock number 021238) is line number one, generated at the University of North Carolina, and currently maintained and distributed by The Jackson Laboratory.

Congenic Mice

When a genetic mutation arises spontaneously or is created in a specific mouse strain and the investigator wants to characterize it on another inbred strain, this can be accomplished by a modification of inbreeding to create what is called a congenic strain (Table 3.9). For example, if you have an albino (white) strain carrying a single gene mutation, specifically a recessive mutation such as hairless on the HRS/J inbred strain (donor strain), and want to move it onto a predominantly C57BL/6J background (host strain), one parent from each strain is mated together to create F1 hybrids (see below). These F1 progeny are intercrossed to create F2 mice, ideally but not always 25% of which will be homozygous for the recessive hairless mutant allele. Only the mutant mice are then crossed back (backcrossed) to C57BL/6J mice to produce the N2 generation. This is repeated 10 times (N10) to create a fully congenic mouse strain [22, 23], with congenic nomenclature permitted to be used at N5. At N10, the strain will be mathematically >99% host background. Unfortunately, there will always be some of the parental strain DNA flanking the gene selected for, known as the congenic interval, and the possibility of trace amounts of donor sequence elsewhere in the genome. The more backcrosses to B6, the smaller this congenic interval should become and the lower the likelihood of traces of donor sequence but the genetic background will never entirely become B6. Creating a group of congenic strains on the same host background controls for phenotypic variability caused by genetic background in comparative or combinatorial assessments of mutation‐induced phenotypes.

This description is only true for recessive mutations that cannot be genotyped and even in those instances, a handful of researchers might go straight to N2 before the intercross, especially if it is a homozygote that cannot be used to breed. Dominant mutations or alleles that can be genotyped do not require the intercross step so can be repeatedly backcrossed at each generation.

Figure 3.7 Using recombinant inbred lines to narrow candidate gene intervals usingWWW.GeneNetwork.org The BXD lines on the left are all homozygous for the hypomorphic Abcc6 allele from DBA/2J. One quantitative trait locus interval has a mixture of DBA/2J (blue) and C57BL/6J (red) DNA. By phenotyping the mice, it is possible to narrow the interval to identify the modifier gene.

      Source: Generated by Dr. Jason Bubier 4 May 2020; WWW.GeneNetwork.org.

Nomenclature for congenic strains begins with a standard abbreviation for the host strain (Table 3.4), followed by a period (.) to indicate congenicity, then the standard abbreviation for the donor strain, then a hyphen, then the allele(s) or congenic interval transferred, then a forward slash (/), and then laboratory code of the investigator in whose laboratory the congenic was completed. For example, a mutation in the sodium channel, voltage‐gated, type VIII, alpha gene (Scn8a^8J) was induced using N‐ethyl‐N‐nitrosourea (ENU) in the C57BL/6J background and was backcrossed onto C3HeB/FeJ in the laboratory of Dr. Wayne Frankel so the strain name is C3Fe.B6‐Scn8a^8J/Frk [24]. In the strain B6.C3‐Pde6b^rd1 Hps4^le/J both the phosphodiesterase 6B, cGMP, rod receptor, β‐polypeptide retinal degeneration one allele (Pde6b^rd1) and HPS4, biogenesis of lysosomal organelles complex 3 subunit 2 light‐ear allele (Hps4^le) were transferred in a single congenic interval from Chromosome 5 of a mutant subline