A simplified method for lysis of single cells in PCR-based preimplantation genetic diagnosis

Complete cell lysis is essential for accurate and efficient polymerase chain reaction (PCR) based-preimplantation genetic diagnosis (PGD). While the methods in current use are efficient, a simpler method would lower the risk of DNA contamination. We therefore sought to develop a simplified technique of single-cell analysis. Single amniocytes and unfertilized oocytes were isolated under a microscope. Twenty cells of each type were individually lysed in buffer containing proteinase K and Tween20, and 20 cells were lysed in buffer containing Tween20 alone. The X chromosome-specific repeat sequence, DXZ1, was subsequently amplified in each cell by PCR. The amplification success rates of individual amniocytes and oocytes lysed in Tween20 were equal to those of the same cell types lysed in Proteinase K+Tween20. The simplified method of cell lysis described here was as efficient as the method currently in use. By reducing the number of manipulations, our method may reduce the possibility of DNA contamination and may be better suited to clinical PGD.