Single-cell PCR on protargol-impregnated euplotid ciliates: a combined approach of morphological and molecular taxonomy

Abstract

Ciliates are considered one of the most diverse protozoa and play significant roles in ecology. For successful taxonomic study of these microscopic eukaryotes, a staining procedure is necessary, due mainly to intrinsic difficulties in recognizing characteristics from living cells. Although molecular taxonomy has been used to resolve the ambiguities associated with traditional morphology-based taxonomy, extraction of genomic DNA from stained ciliate cells is not available yet. In the present study, we describe a method to extract genomic DNA from a single protargol-impregnated euplotid cell. By using HgCl(2) as a fixative and modulating the exposure time of bleach solution in the protargol impregnation, high-quality genomic DNA can successfully be extracted from a stained single cell with minimal loss of morphological integrity. This technique will contribute to the effectiveness of combined approaches of molecular and morphological taxonomy from single ciliate cells.