| 18844099 |
Long MJ, Song F, Qu YJ, Meng Y, Wang H, Jin YW, Huang SZ: [Quantitative analysis of SMN1 and SMN2 genes based on DHPLC: a reliable method for detection of non-homozygous patients with spinal muscular atrophy]. Zhonghua Yi Xue Za Zhi. 2008 May 13;88(18):1259-63.
OBJECTIVE: To develop a rapid and reliable approach for testing the copy number of survival motor neuron (SMN) gene and analyze the compound heterozygous deletions of SMN1 gene. METHODS: Peripheral blood samples were collected from 38 non-homozygous deletion pediatric patients with SMA, 30 homozygous deletion patients with SMA, and 35 un-related healthy persons. SMN1 and SMN2 genes were amplified separately with allele-specific PCR (AS-PCR). Meanwhile, two irrelevant genes were amplified as internal quality control respectively. The copy numbers of SMN1 and SMN2 were determined by denaturing high-performance liquid chromatography (DHPLC). RESULTS: (1) A protocol combining multiplex allele-specific PCR and DHPLC was developed to separate SMN1 and SMN2 and to determine the copy numbers of them. The copy numbers of SMN1 and SMN2 varied from 1 to 4 and a clear-cut differentiation among the different copy number ranges could be observed for the two genes. (2) One single copy of SMN1 were detected in 20 of the 38 non-homozygous deletion patients with SMA (52.6%). Heterozygous deletions were determined in these 20 patients. Two copies of SMN2 were detected in 15 of the 20 patients with one copy of SMN1 (75.0%, 15/20). Other 5 of the 20 patients were with 3 copies of SMN2 (25.0%, 5/20). (3) One single copy of SMN1 was detected in 24 of the 30 (80%) parents of SMA patients with homozygous deletion. CONCLUSION: SMN copy number can be rapidly and reliably determined by the method of multiplex AS-PCR combined with DHPLC. |
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