[Limitation of PCR-RFLP method for the detection of genetic mutations in spinal muscular atrophy].
OBJECTIVE
To explore the applicability and limitation of PCR-restriction fragment length polymorphism (PCR-RFLP) method for genetic diagnosis of spinal muscular atrophy (SMA).


METHODS
PCR-RFLP was applied to detect potential deletion in exons 7 and 8 of SMN1 gene in 935 suspected cases with SMA. Multiplex ligation-dependent probe amplification(MLPA) was carried out to analyze dosage alteration of SMN1 gene in 339 of such cases. To confirm the accuracy of PCR-RFLP method for homozygous and heterozygous deletions detection, the consistency of PCR-RFLP and MLPA results were assessed with a Pearson Chi-square test.


RESULTS
Homozygous deletion of exon 7 of SMN1 was detected in 590 suspected cases. The rate of diagnosis was therefore 63.1% (590/935). For the 339 suspected cases, PCR-RFLP and MLPA respectively identified 194 and 196 homozygous deletions in the exon 7 of SMN1 gene, suggesting a good consistency (98.9%)(Chi-square = 0.2, P = 0.88). However, only 4 of 339 cases was found to carry a heterozygous deletion of SMN1 exon 7 by PCR-RFLP, in contrast with 17 detected by MLPA. The consistency only reached 23.5%, for which statistical significance was detected (Chi-square = 8.29, P< 0.01).


CONCLUSIONS
Although PCR-RFLP is a simple, specific and efficient method for SMA diagnosis, it has obvious limitation for the diagnosis of  5%-10% SMA patients who have carried a compound heterozygous mutation.
Department of Medical Genetics, Capital Institute of Pediatrics, Beijing 100020, P. R. China.
Yu-wei
YW
Jin
Yu-jin
YJ
Qu
Hong
H
Wang
Jin-li
JL
Bai
Fang
F
Song
D005091:Exons; D006801:Humans; D009134:Muscular Atrophy, Spinal / Q000235:genetics*; D009154:Mutation; D016133:Polymerase Chain Reaction / Q000379:methods*; D012150:Polymorphism, Restriction Fragment Length