Quantitative analysis of SMN1 and SMN2 genes based on DHPLC: A highly efficient and reliable carrier-screening test
Journal
Human Mutation
Journal Volume
25
Journal Issue
5
Pages
460-467
Date Issued
2005
Author(s)
Su Y.-N.
Hung C.-C.
Li H.
Chang M.-C.
Hsieh W.-S.
Hsu S.-M.
Abstract
Autosomal recessive spinal muscular atrophy (SMA) is a common, fatal neuromuscular disease caused by homozygous absence of the SMN1 gene in approximately 94% of patients. However, a highly homologous SMN2 gene exists in the same chromosome interval, centromeric to SMN1, and hampers detection of SMN1. We present a new, rapid, simple, and highly reliable method for detecting the SMN1 deletion/conversion and for determining the copy numbers of the SMN1 and SMN2 genes by DHPLC. We analyzed SMN1/SMN2 gene exon 7 deletion/conversion by DHPLC. A total of 25 patients with spinal muscular atrophy lacking the SMN1 gene as well as 309 control individuals from the general population and the family members of patients with SMA were analyzed. By DHPLC analysis, we could detect the SMA-affected cases efficiently just by recognizing an SMN2-only peak. Furthermore, after specific primer amplification and adjustment of the oven temperature, all of the SMA carriers with an SMN1/SMN2 ratio not equal to 1 could be identified unambiguously by this simple and efficient detection system. To calculate the total SMN1/SMN2 gene dosages further, we developed a specific multiplex competitive PCR protocol by simultaneously amplifying the CYBB gene (X-linked), the KRIT1 gene (on chromosome arm 7q), and the SMN1/SMN2 gene ratio by DHPLC. By applying this technique, we could successfully designate all of the genotypes with different SMN1/SMN2 gene copy numbers, including equal and unequal amounts of SMN1 and SMN2. We demonstrated that DHPLC is a fast and reliable tool for detection of carriers of SMA. ? 2005 Wiley-Liss, Inc.
SDGs
Other Subjects
survival motor neuron protein; survival motor neuron protein 1; survival motor neuron protein 2; unclassified drug; article; clinical article; controlled study; CYBB gene; denaturing high performance liquid chromatography; exon; gene; gene amplification; gene conversion; gene deletion; gene dosage; human; krit1 gene; nucleotide sequence; polymerase chain reaction; population research; priority journal; quantitative analysis; screening test; spinal muscular atrophy; Chromatography, High Pressure Liquid; Cyclic AMP Response Element-Binding Protein; Female; Gene Dosage; Genetic Screening; Heterozygote Detection; Humans; Male; Molecular Sequence Data; Muscular Atrophy, Spinal; Nerve Tissue Proteins; Pedigree; RNA-Binding Proteins
Type
journal article