Quality controls for cell cultures: identification of interspecies cross-contamination by PCR-RFLP analysis of the cytochrome b gene

Cross-contaminations of a cell line with cells of different species represent a potential risk in laboratories handling human and animal cells. Therefore, it is necessary to control such contaminations. Tests based on mitochondrial DNA (mtDNA) are used in forensic analysis, phylogenetic studies and in food authentication. However, the use of mtDNA in quality controls of cell cultures is recent. Mitochondrial sequence differences of closely related animal species are five- to tenfold higher than those of nuclear genes. On the contrary, intraspecies variation in mitochondrial sequences is low in most animal species. Moreover, each cell contains 100–10.000 mitochondrial genomes. The amount of mtDNA is greater than nuclear DNA, so that mtDNA can be analyzed also from small or partially degraded samples. In the present study, a method based on a PCR-Restriction Fragment Length Polymorphism (RFLP) analysis of the mitochondrial cytochrome b gene was used (2). This gene has some stable sequences which are recognized from universal primers and some variable sequences used for animal species identification by PCR-RFLP method.