Single-cell genetic analysis includes single-cell DNA and FISH analysis, which have become important tools for application of PGD in assisted reproduction and genetic services, providing a practical option for couples at genetic risk to avoid the birth of an affected offspring and have a healthy child of their own.
Because PGD for single-gene disorders is based on single-cell genetic analysis, its accuracy depends largely on the limitations of single-cell DNA analysis, which may potentially cause misdiagno-sis. One of the key contributors to misdiagno-sis is the phenomenon of preferential amplification, also known as allele-specific amplification failure (allele drop out, ADO), requiring the application ofspecial protocols to ensure the highest ADO detection rate [29, 42, 43]. A few previously reported misdiagnoses, involving PGD for beta-thalassemia, myotonic dystrophy (DM), fragile-X syndrome (FMR1) and cystic fibrosis (CF), might have been due to this phenomenon, which has not initially been fully realized [21, 23, 28, 44].
It has been demonstrated that ADO rates in single cells might be different for different types of heterozygous cells . The ADO rate may exceed 20% in blastomeres, compared with the ADO rate in single fibroblasts and PB1, which was shown to be under 10%. A high rate of ADO in blastomeres may lead to an obvious misdiagnosis, especially in compound heterozygous embryos. As mentioned, most misdiagnoses, especially those at the initial stage of application of PGD for single-gene disorders, were in the cases of blastomere biopsy from apparently compound heterozygous embryos.
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