Genetic Diagnosis of Spinal Muscular Atrophy Using MLPA

Objective: To perform genetic diagnosis for 30 suspected patients with spinal muscular atrophy (SMA) using multiplex ligation dependent probe amplification (MLPA) and PCR-restriction fragment length polymorphism (PCR-RFLP), and compare the results of the two methods. Methods: The genomic DNA was isolated using salting-out method from peripheral blood of each subject from 30 families.Exon 7 and 8 of SMN genes were amplified by allele specific PCR. The PCR products were digested with DralⅠand DralⅠdigested PCR products. Simultaneously, the DNA samples were analyzed by SALSA MLPA Kit P021. Results: Both PCR-RFLP and MLPA analysis showed the same that 22 patients with exon 7 and 8 homozygous deletion, and 2 patients with only exon 7 homozygous deletion of SMN1. The other 6 cases presented no homozygous deletion by PCR-RFLP, but one child and two mothers of them were detected heterozygous by MLPA. Also MLPA analysis found three 2+0carriers from 3 families. The data also showed that the SMN2 copy numbers were mainly 4 or 5 in SMA patients, while the carriers and the normal individuals were 2 or 3 and 1 or 2 copies respectively. There was clear statistical significance in the group of patient -carrier and group of patient-normal individuals (P 0.01). The carrier -normal group appeared no statistical significance (P 0.05). Conclusion: Compared with PCR-RFLP, MLPA is more convenient, precise, high-effective, and it can accurately quantitative analysis of SMN1 and SMN2. MLPA is a kind of technique of genetic diagnosis for common genetic disease.