Gene dosage analysis of proximal spinal muscular atrophy carriers using real-time PCR.

BACKGROUND Autosomal recessive spinal muscular atrophy is a disease resulting from homozygous absence of SMN1 gene in approximately 94% of SMA patients. To identify patients who retained a single SMN1 copy, SMN1 dosage analysis was performed by quantitative Real-time PCR using SYBR green dye. SMN1 dosage analysis results were utilized to identify carriers before offering prenatal diagnosis. METHOD Carrier testing was performed for 150 individuals. Copy number of the SMN1 gene was determined by the comparative threshold cycle (Ct) method and human serum albumin gene was used as a reference. RESULT Analysis of 150 DNA samples with quantitative PCR determined the number of SMN1 gene copies. Of these, 50 (33.33%) cases had one SMN1 gene copy, 87 (58%) had two copies and 13 (8.66%) did not have any copies of SMN1. The homozygous SMN1 deletion ratio was 0.00 and deletion of one copy of SMN1 gene ratio ranged from 0.3 to 0.58. CONCLUSION This report demonstrates modification of risk estimation for the diagnosis and detection of SMA carriers by accurate determination of SMN1 copy number. SMN1 copy number analysis is an important parameter for identification of couples at risk of having children affected with SMA. It also reduces unwarranted prenatal diagnosis for SMA. Furthermore, the dosage analysis might be useful for the counseling of clinically suspected SMA patients with negative diagnostic SMA tests.

[1]  E. Tizzano,et al.  Carrier frequency of SMA by quantitative analysis of the SMN1 deletion in the Iranian population , 2010, European journal of neurology.

[2]  K. Kahrizi,et al.  Deletions in the survival motor neuron gene in Iranian patients with spinal muscular atrophy. , 2009, Annals of the Academy of Medicine, Singapore.

[3]  صادق ولیان بروجنی,et al.  Molecular analysis and pr enatal diagnosis of spinal muscular atrophy in Iranian population: association of neural apoptosis inhibitory protein (NAIP) deletions with severity of the disease , 2008 .

[4]  D. Farhud,et al.  Molecular analysis of the SMN1 and NAIP genes in Iranian patients with spinal muscular atrophy. , 2007, Annals of the Academy of Medicine, Singapore.

[5]  Hyun-Seok Jin,et al.  Quantitative Analysis of SMN1 Gene and Estimation of SMN1 Deletion Carrier Frequency in Korean Population based on Real-Time PCR , 2004, Journal of Korean medical science.

[6]  B. Wirth,et al.  Best practice guidelines for molecular analysis in spinal muscular atrophy , 2001, European Journal of Human Genetics.

[7]  B. Wirth.  An update of the mutation spectrum of the survival motor neuron gene (SMN1) in autosomal recessive spinal muscular atrophy (SMA) , 2000, Human mutation.

[8]  J. McPherson,et al.  A single nucleotide difference that alters splicing patterns distinguishes the SMA gene SMN1 from the copy gene SMN2. , 1999, Human molecular genetics.

[9]  J. Mendell,et al.  Identification of proximal spinal muscular atrophy carriers and patients by analysis of SMNT and SMNC gene copy number. , 1997, American journal of human genetics.

[10]  C. Ponting,et al.  Missense mutation clustering in the survival motor neuron gene: a role for a conserved tyrosine and glycine rich region of the protein in RNA metabolism? , 1997, Human molecular genetics.

[11]  L. Kunkel,et al.  A multicopy transcription-repair gene, BTF2p44, maps to the SMA region and demonstrates SMA associated deletions. , 1997, Human molecular genetics.

[12]  J. Mendell,et al.  Deletion and conversion in spinal muscular atrophy patients: Is there a relationship to severity? , 1997, Annals of neurology.

[13]  J. Melki,et al.  Frameshift mutation in the survival motor neuron gene in a severe case of SMA type I. , 1996, Human molecular genetics.

[14]  E. Velasco,et al.  Molecular analysis of the SMN and NAIP genes in Spanish spinal muscular atrophy (SMA) families and correlation between number of copies of cBCD541 and SMA phenotype. , 1996, Human molecular genetics.

[15]  A. Munnich,et al.  A frame–shift deletion in the survival motor neuron gene in Spanish spinal muscular atrophy patients , 1995, Nature Genetics.

[16]  B. Wirth,et al.  Molecular analysis of candidate genes on chromosome 5q13 in autosomal recessive spinal muscular atrophy: evidence of homozygous deletions of the SMN gene in unaffected individuals. , 1995, Human molecular genetics.

[17]  B. Dallapiccola,et al.  Survival motor neuron gene transcript analysis in muscles from spinal muscular atrophy patients. , 1995, Biochemical and biophysical research communications.

[18]  J. Osinga,et al.  PCR-based DNA test to confirm clinical diagnosis of autosomal recessive spinal muscular atrophy , 1995, The Lancet.

[19]  K. Davies,et al.  Deletions in the survival motor neuron gene on 5q13 in autosomal recessive spinal muscular atrophy. , 1995, Human molecular genetics.

[20]  T. Crawford,et al.  The gene for neuronal apoptosis inhibitory protein is partially deleted in individuals with spinal muscular atrophy , 1995, Cell.

[21]  J. Weissenbach,et al.  Identification and characterization of a spinal muscular atrophy-determining gene , 1995, Cell.

[22]  D. Le Paslier,et al.  De novo and inherited deletions of the 5q13 region in spinal muscular atrophies. , 1994, Science.

[23]  J. Pearn.  CLASSIFICATION OF SPINAL MUSCULAR ATROPHIES , 1980, Lancet.

[24]  A. Munnich,et al.  The gene encoding p44, a subunit of the transcription factor TFIIH, is involved in large-scale deletions associated with Werdnig-Hoffmann disease. , 1997, American journal of human genetics.