Korean J Fertil Steril.  2005 Mar;32(1):17-26.

Efficacy of Duplex-nested PCR and Fluorescent PCR in the Preimplantation Genetic Diagnosis for Duchenne Muscular Dystrophy

• Lee HS
• Choi HW
• Lim CK
• Park SY
• Kim JY
• Koong MK
• Jun JH
• Kang IS

Abstract

OBJECTIVE
Preimplantation genetic diagnosis (PGD) is reserved for couples with a risk of transmitting a serious and incurable disease, and hence avoids the undesirable therapeutic abortion. In this study, we evaluated the efficacy of PGD for Duchenne muscular dystrophy (DMD) cases by the fluorescent PCR with polymorphic linked markers and the conventional duplex-nested PCR methods.
METHODS
Biopsy of one or two blastomeres was done from the embryos fertilized by ICSI on the third day after fertilization. We performed two cases of PGD-DMD by the duplex-nested PCR for the causative mutation loci and the SRY gene on Y chromosome. The triplex fluorescent PCR for the mutation loci, the SRY gene and the polymorphic microsatellite marker on X chromosome was applied for two cases of PGD-DMD.
RESULTS
By the duplex-nested PCR, successful diagnosis rate was 95.5% (21/22), but we could not discriminate the female embryos whether normal or carrier in this X-linked recessive disease. However, the triplex fluorescent PCR method showed 100% (27/27) of successful diagnosis rate, and all female embryos (n=17) were distinguished normal (n=10) from carrier (n=7) embryos. Unaffected and normal embryos were transferred into mother's uterus after diagnosis. A healthy normal male was achieved after PGD with the duplex-nested PCR method and a twin, a male and a female, were delivered with triplex fluorescent PCR method. The normality of dystrophin gene was confirmed by amniocentesis and postnatal genetic analysis in all offsprings.
CONCLUSION
The fluorescent PCR with polymorphic marker might be useful in improving the specificity and reliability of PGD for single gene disorders.

Keyword

Preimplantation genetic diagnosis (PGD); Duchenne muscular dystrophy (DMD); Duplexnested PCR; Fluorescent PCR; Polymorphic marker

MeSH Terms

Abortion, Therapeutic
Amniocentesis
Biopsy
Blastomeres
Diagnosis
Dystrophin
Embryonic Structures
Family Characteristics
Female
Fertilization
Genes, sry
Humans
Male
Microsatellite Repeats
Muscular Dystrophy, Duchenne*
Polymerase Chain Reaction*
Pregnancy
Preimplantation Diagnosis*
Prostaglandins D
Sensitivity and Specificity
Sperm Injections, Intracytoplasmic
Twins
Uterus
X Chromosome
Y Chromosome
Dystrophin
Prostaglandins D