Simple and Easy to Perform Preimplantation Genetic Diagnosis for β‑thalassemia Major Using Combination of Conventional and Fluorescent Polymerase Chain Reaction

Thalassemias are the most common monogenic disorders in many countries throughout the world. The best practice to control the prevalence of the disease is prenatal diagnosis (PND) services. Extensive practicing of PND proved effective in reducing new cases but on the other side of this success high abortion rate is hided, which ethically unfair and for many couples, especially with a previous experience of a therapeutic abortion, or moral concerns, is not a suitable choice. Preimplantation genetic diagnosis (PGD) is a strong alternative to conventional PND. At present PGD is the only abortion free fetal diagnostic process. Considering the fact that there are more than 6000 single gene disorders affecting approximately 1 in 300 live‑births, the medical need for PGD services is significant.

In the present study development of a PGD protocol for a thalassemia trait couple using nested multiplex fluorescent polymerase chain reaction (PCR) for the combination of polymorphic linked short tandem repeat (STR) markers and thalassemia mutations is described. Restriction fragment length polymorphism used to discriminate between wild and mutated alleles.

In PGD clinical cycle, paternal and maternal alleles for D11S988 and D11S1338 STR markers were segregated as it was expected. PCR product for IVSII‑1 mutation was subsequently digested with BtscI restriction enzyme to differentiate normal allele from the mutant allele. The mother’s mutation, being a comparatively large deletion, was detectable through size differences on agarose gel.

The optimized single cell protocol developed and evaluated in this study is a feasible approach for preimplantation diagnosis of β‑thalassemia in our patients.