Risks and pitfalls

The study of specific DNA sequences in a single blastomere is a substantial challenge as many problems may compromise both the accuracy and reliability of the genetic diagnosis. For the detection of single gene defects, the prevalent method of diagnosis is PCR (polymerase chain reaction) allowing the exponential amplification of short DNA sequences, which are subsequently detected and analysed.

Since the first PCR-based PGD cases, genetic analysis for single gene disorders has evolved considerably. However, major difficulties are still encountered in single cell analysis because of the minute amount of DNA available :

- DNA contamination of the sample with extraneous non-embryonic DNA,

- Amplification failure,

- Allele drop out (ADO), which is defined as the random non-amplification or non-detection of one of the alleles present in a heterozygous sample,

- Preferential amplification of one allele.

All couples have to be aware of these technical limitations as they may lead to absence of diagnosis or to potential misdiagnosis.

Amplification failure is highly undesirable, as no diagnosis is obtained and no embryo may be transferred, but ADO is much more dangerous as it can lead to an unacceptable misdiagnosis with the transfer of an affected embryo. In autosomal recessive disorders, ADO of the unaffected allele would lead to the non transfer of an unaffected embryo. ADO is particularly of concern in PGD for autosomal dominant disorders where ADO of the affected allele could lead to the transfer of an affected embryo. Various strategies have been developed to minimize and detect this phenomenon.