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[Objective]: using real-time quantitative PCR method, to establish a stable, fast, and accurate survival motor neuron gene (survival motor neuron SMN1) quantitative methods, and used in spinal muscular atrophy (spinal muscular atrophy, SMA ) carriers of the screening, as well as the evaluation of the SMA carrier screening for genetic diagnosis and prenatal diagnosis of SMA. [Method]: In the TaqMan technology, the use of the specificity of the MGB probe method, the simultaneous selection factor Ⅷ exon 3 homogenization correction of the initial concentration of sample DNA as a reference gene. The establishment of standards, the standards and the test sample at the same time in charge of SMN1 gene was amplified Ⅷ factor to optimize the reaction conditions, the design of standardized testing procedures, serial dilutions of standard building the SMN1 gene Ⅷ factor standard curve. Treated by the relationship between the Ct value and standard curve of the sample measurements the SMN1 and VIII of the reaction starting template for quantitative analysis, to thereby obtain sample genomic SMN1 copy number. In order to verify the reproducibility and stability of the RQ-PCR, respectively, for men and women carriers, male and female normal control four different sample batch and batch comparison; same time, 48 cases of of SMA patients diagnosed by RFLP-PCR method and 20 cases were detected by linkage analysis to determine the affirmative carriers to verify the specificity and reliability of the technology. In this study, by detecting the SMN1 copy number of the general population of 200 cases, 26 cases of parents of SMA patients and 16 fetuses SMA carrier screening. the grant and the copy number of interval [1.70-2.18] and [1.68-1.98]; specificity and reliability experiment, 48 cases of the SMA patient samples SMN1 copy number 0,20 cases certainly carriers copy number range of 0.71-1.15, average copy number of 0.90 ± 0.135. 2.200 cases of the general population in the four cases containing a copy of the SMN1 gene carriers, its copy number range of 0.75-1.05, average copy number was 0.88 ± 0.162 consistent, and of SMN1 certainly carriers range: 194 cases of copy number range of 1.60-2.18, average copy number of non-carriers: the remaining two cases SMN1 gene / Ⅷ factor ratio is 1.88 ± 0.156, 3.09 and 3.22, respectively, with an average copy number of 3.16 ± 0.092, containing three copies of SMN1 gene carriers. 3. Parents sample of 26 cases, 25 cases of copy number range 0.61-1.16, average copy number was 0.91 ± 0.163,96.1% for carriers: only 1 case of copy number 1.82,3.9% in line with the scope of the dual copy. 16 fetuses sample, including 10 cases of copy number range of 1.62-2.11, average copy number of 1.88 ± 0.147, a non-carriers, the remaining six cases, copy number interval 0.73-1.10, average copy number of 0.90 ± 0.194, SMN1 gene deletion carriers. [Conclusion]: real-time quantitative PCR can be used for quantitative SMN1 gene detection, accurate and reliable, simple, practical, and automated scale detection system. The SMN1 gene 2.SMA carriers and non-carriers of the copy number Interval Different SMN1 gene copy number can be used for screening SMA carrier quantitative analysis. Applications SMA carrier screening for SMA genetic counseling, genetic risk assessment, genetic diagnosis has important scientific value and clinical significance, and provide an important basis for the genetic diagnosis and prenatal diagnosis of SMA.
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