Transgenic Research

, Volume 22, Issue 6, pp 1265–1272

Combining sperm plug genotyping and coat color chimerism predicts germline transmission

  • Angus Yiu-Fai Lee
  • Kristin Evans
  • Brandon Willis
  • K. C. Kent Lloyd
Brief Communication

DOI: 10.1007/s11248-013-9731-6

Cite this article as:
Lee, A.YF., Evans, K., Willis, B. et al. Transgenic Res (2013) 22: 1265. doi:10.1007/s11248-013-9731-6

Abstract

There has been a significant increase in the use of C57BL/6N-derived ES cells for the production of gene knockout mice. However, the potential for germline transmission (GLT) from chimeras on this genetic background has been observed to be highly variable. Using coat color as an indicator of somatic chimerism to infer the extent of chimeric contribution to the germ cell population, even highly agouti C57BL/6N-derived chimeras can fail to achieve GLT. We investigated the extent to which quantitative PCR genotyping for a marker gene expressed in gene targeted ES cells can be performed on DNA extracted from sperm present in copulatory plugs to determine the contribution of ES cells to the germ cells. We found that an objective assessment of sperm DNA from copulatory plugs combined with a subjective assessment of coat color chimerism can be used to accurately inform the selection of chimeras for breeding that are likely to achieve GLT. These results indicate that, compared to random selection of chimeras, including an analysis of copulatory plugs to set chimeras for breeding can help to reduce costs, minimize time, and facilitate research for projects requiring the production, selection, breeding, and testing of chimeras to generate gene-targeted mice.

Keywords

ES cells C57BL/6N Chimera Germline transmission Copulatory plug Quantitative PCR Gene knockout 

Copyright information

© Springer Science+Business Media Dordrecht 2013

Authors and Affiliations

  • Angus Yiu-Fai Lee
    • 1
  • Kristin Evans
    • 1
  • Brandon Willis
    • 1
  • K. C. Kent Lloyd
    • 1
  1. 1.Mouse Biology ProgramUniversity of California, DavisDavisUSA