Problems with Single Cell PCR

Single-cell PCR is technically demanding, with high risks of contamination and ADO. Contamination can occur from the operator's DNA or carryover of previous PCR products. This type of contamination can be avoided by working in sterile manners, using different pre- and post-PCR rooms and materials dedicated only for the setup of single-cell PCR. Another source of contamination is genomic DNA from the patients, e.g., maternal cumulus cells or sperms sticking to the zona pellucida accidentally sampled during biopsy procedures. To avoid the risk of paternal contamination, ICSI should be used to achieve fertilization in all cases of PCR-based PGD. Al-lele dropout is a phenomenon observed when one of the examined alleles fails to amplify during the PCR reaction. This can result in diagnostic errors, especially for autosomal dominant disorders, as ADO of the mutated allele would wrongly diagnose an affected embryo as healthy.

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