Mouse tissue culture models of unstable triplet repeats.
Once into the expanded disease-associated range, trinucleotide repeat alleles become dramatically unstable in the germline and in somatic cells. The molecular mechanism(s) that underlie this unique form of dynamic mutation are poorly understood. Numerous transgenic mouse models of unstable trinucleotide repeats, which reconstitute the dynamic nature of somatic mosaicism observed in humans, have been generated. Given their easy accessibility, tissues from these mice can be collected to establish homogenous cell culture models of trinucleotide repeat dynamics. This chapter describes how such cultures can be established and maintained. Such in vitro systems may be useful to study relevant biological questions concerning fundamental triplet repeat metabolism. In particular, monitoring of repeat stability in cells growing under controlled conditions could help to clarify the relationship among the accumulation of repeat length variation, cell division rates, and DNA replication.
A microtechnique (modified Takatsy) is described which can be applied to complement fixation, hemagglutination, hemagglutination inhibition and metabolic inhibition tests. The system permits an 8-fold saving of reagents and rapid performance of microdilutions. The specific methods and modifications of equipment necessary to obtain reliable results are presented in detail. Comparative data obtained with the micro- and standard systems establish the reliability and validity of the microsystem.