Chromosome Counts for ES Cells - Testing for Euploid Clones

1. Why count chromosomes in gene targeted ES cells?

It is recognized that not all cell lines in culture have normal chromosome complements. Numerous groups have observed that aneuploid ES cell clones do not form germline ES cell-mouse chimeras (exception: germline transmission from X:O ES cells can occur through female chimeras). We routinely detect aneuploid ES cell clones in our experiments, in gene trap clones imported from international consortia, and in clones generated independently by other laboratories. Deriving and breeding ES cell chimeras from aneuploid ES cells is expensive with respect to animals used, time, and money. The simple chromosome counting method below can improve the efficiency of germline transmission in any experiment. We count 20 spreads per ES clone in our Transgenic Core Facility. We microinject ES cell clones in which 60% or more of the spreads contain 40 chromosomes. Note that simply because most of the chromosome spreads in an ES cell clone are euploid is no guarantee that the clone will go germline.

2. Why not karyotype chromosomes in gene targeted ES cells?

A karyotype shows and identifies each chromosome in a spread. This identification of the chromosomes in multiple spreads is a time consuming process.  Commercial services for karyotyping are available, but can add dramatically to the cost of a gene targeting project. Although some ES cell lines with 40 chromosomes will be aneuploid because of chromosome duplications and deletions, the proportion of ES cell clones with euploid chromosome counts that go germline is so high that there is no real advantage to screening the cells by karyotype analysis.

Download Chromosome Counting Protocol

 
 
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