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May 29, 2009

Shortening the generation of generations

Immature mice father faster medical research via an assisted reproduction technique that speeds up the production of mouse models

Figure 1: Germ cells (dotted circle, right) collected from a 22-day old male (black mouse) can be used in high-speed breeding of genetically engineered mouse models.

Japanese researchers have developed a breeding strategy to produce strains of genetically modified mice with precise genetic backgrounds in about six months. This compares to three to four years using conventional breeding strategies and is more than twice as fast as now possible with the latest ‘speed’ methods. The new strategy, based on existing assisted reproduction techniques, should accelerate the genetic analysis integral to biomedical research using mouse models.

After decades of experience in engineering laboratory mice, geneticists can now alter mouse genomes with the precision of a single nucleotide change, and create mice with specific genetic modifications. The biological function of genes, however, can vary depending on their genetic background, and the easiest genomes to alter often result in unexpected or irreproducible individuals. So an engineered change must typically be back-crossed into a standard inbred strain.

Using classical breeding techniques it takes 10 generations, or three to four years, to produce strains that differ by less than 1%, including the desired genetic change. This is time consuming and expensive. Recently, employing markers associated with particular groups of genes, this process has been accelerated to four to five generations or one to two years.

Geneticists then began to wonder whether the process could be accelerated further by using hormones to mature and ovulate the eggs of immature female mice to shorten the generation time. But the number of eggs able to be treated this way is limited and the response to hormones is variable.

So Atsuo Ogura and colleagues from the RIKEN BioResource Center and the universities of Tokyo and Tsukuba tried the reverse. In a recent paper published in the journal PLoS ONE1, they detail how they used a technique known as round spermatid injection (ROSI) to extract the first wave of round spermatids—the earliest stage of maturing spermatozoa—from immature male mice and inject them into the eggs of mature females producing healthy offspring (Fig. 1).

The optimum age of male mice used was 22 days old, shortening the generation time to about 40 days, and the establishment of genetically specific strains to less than 200 days. They have applied their technique to generate separate strains of genetically modified mice: transgenic mice; ‘knock-in’ mice with a DNA sequence added; and mutant mice that were produced using the chemical N-ethyl-N-nitrosourea. “Our next goal is to make the technique easier,” Ogura says, “in the hope that it will become more popular.”

References

  1. Ogonuki, N., Inoue, K., Hirose, M., Miura, I., Mochida, K., Sato, T., Mise, N., Mekada, K., Yoshiki, A., Abe, K., Kurihara, H., Wakana, S. & Ogura, A. A high-speed congenic strategy using first-wave male germ cells. PLoS ONE 4 (3), e4943 (2009). |  | (Link)