Zpráva z pracovní cesty - Ploen (Německo)

From 4th to 26th of October, Oleksiy Yanchukov (post-doc) visited a Max-Plank Institute of Evolutionary Biology in Ploen, Germany.

There he conducted a series of laboratory experiments in order to genotype the genomic DNA samples of the house mice from the Czech-Bavarian hybrid zone for gene copy-number (CN) variation. Using the QX-200 droplet digital PCR (ddPCR) machine (Bio-Rad), he attempted to quantify the signals form the reference single-copy gene(s) and the target gene(s) with unknown copy-number, based on the difference in ampicon lengths, using a single colour detection with Eva-Green DNA dye. After a series of experiments, it was possible to distinguish the samples with elevated gene copy number, however, the precision of such quantification can be rather poor due to the insufficient separation of signals from the reference and the target genes. He concluded that the single colour multiplexing could be used a cheap and quick method to scan for the elevated CN among many samples. To precisely determine the copy number, a smaller subset of samples with suspected CN differences can be amplified on the same ddPCR instrument using the specific fluorescent probes (a more expensive). Alternatively, the specific probes may be used when the precision is important and samples are not numerous. In the future visits to Ploen, a combination of both methods will be used.