Watch your Back!

A whistle-stop tour of background strains

The ancestry of the laboratory mouse involves centuries of ‘fancy mouse’ breeding, the importing of many strains from the Far East to Europe and North America by Victorian collectors and the innovative breeding schemes of a woman named Abbie Lathrop from Illinois who bred the first inbred strains. A hundred years on, it is estimated that the number of genetically distinct inbred strains housed in mouse laboratories across the world numbers into the thousands. The official guidelines for nomenclature and ancestry of many is registered at the Mouse Genome Informatics database (MGI).

Inbred strain- a strain which has been derived from 20 or more generations of brother-sister mating (and is therefore homozygous throughout almost all of its genome).

The game-changing transgenic technologies of the 1980s have meant that the manipulation of a single gene of the mouse genome has become routine. The excitement in generating  new  disease models for drug development and novel genetically altered mouse strains for investigating fundamental biological processes has been dampened, however, by reports of non-translatability and irreproducible data. There are many variables in in vivo science which we could point to as being the source of this, including environmental effects, assay conditions and the interpretation of analysis. However, there is no doubt that the poor characterisation or description of the genomes which form the context of published gene manipulations are a major contributory factor.

Genetic background refers to the genetic make-up of the mouse strain(s) which have been used to produce the genetically altered (GA) mouse. This is usually one or two inbred strains and their subsequent crosses.

When generating new GA mouse strains, usually wild type inbred strains or F1 crosses between two strains are used. The phenotype resulting from genetic manipulations will not only be a consequence of the gene which is being changed (e.g. knocked out or mutated) but also a result of the complex interactions of the other genes within the genome of the inbred strains used. Recent years have seen the publication of many papers documenting the diversity of the genomes of inbred mice1 and the phenotypic consequences of these different genetic combinations. Moreover the sequencing of 17 inbred strains in 20112 and their subsequent annotation led to the realisation that the genomes of inbred strains harbour many differences in both coding and non-coding DNA sequences and that these undoubtedly have profound effects on phenotypes. The extent of these differences has both surprised and shocked researchers with over 50 million SNPs being identified between these 17 common laboratory strains.

SNP- single nucleotide polymorphisms – a DNA sequence variation in which a single nucleotide differs when two genomes are compared.

Single gene mutations harboured by inbred strains are conceptually easy to understand in terms of their potential relevance to the phenotypes of mutant strains. For example, the presence of Pde6rd1  and Cdhahl alleles in C3H and C57BL/6 strains respectively render these mice either blind or deaf during their lives. Obviously these strains are inappropriate for studies of either vision or hearing. However the mechanisms of complex gene traits are much more difficult to describe. Mouse lines maintained on mixed or undefined background are much more susceptible to phenotypic variation as the mixes of genes alter from generation to generation and are often virtually impossible to recapitulate.

So how can you ensure that the genetic background of the mouse line you are investigating does not contribute to generating unrepeatable or misleading data? Here’s the top ten tips for watching your back(ground)!

  1. Generate new GA mouse lines on defined, genetically controlled inbred backgrounds.
  2. Avoid transgenesis on F1s or outbred strains when in vivo phenotyping is your aim.
  3. Where the background is unknown (or you have used F1s or outbreds), breed the mice to congenicity (ten generations of backcrossing) onto a known strain of choice.
  4. When the option to backcross is not available (e.g. time or resources not available), control carefully with mice of an appropriate genetic backgrounds such as littermates (although they will never be the same if this is a mixed line).
  5. Be prepared for phenotypes to change as you backcross.
  6. When supplying lines to collaborators, make sure they are exported with full details of the ancestral breeding schemes and strains.
  7. When importing lines from collaborators, ask for full details including the history of their inbred stocks and how many generations they have been bred at their establishment.
  8. If maintaining an inbred strain, ensure that you return to the ancestral stock and restock periodically (every ten generations).
  9. Search public databases (see below) which contain many details of phenotypes, including difference for the same genetic alteration which can be potentially be attributed to differences in genetic background in preparation for working on a new gene/strain.
  10. In summary- be aware! Know what background you are working with, whether it is likely to change with time and plan appropriate controls.

Public GA databases

References

  1. Frazer et al 2007 (Nature 480:1050)
  2. Keane et al 2011 (Nature 477:289)

By Sara Wells, MRC Harwell

Last updated: November 2014