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Objective: To study homologous genes using SYBR Green I quantitative PCR (homologous gene quantitative PCR, HGQ-PCR) diagnostic 21 - trisomy syndrome (Down syndrome, DS), seeking a simple, fast, accurate, safe and cost-effective diagnosis of DS Methods. Method: 1. Establishing method: SYBR Green I HGQ-PCR: Select a pair of primers to simultaneously amplify the long arm of chromosome 21 in human liver-type phosphofructokinase (the human liver-type phosphofructokinase located on chromosome 21, PFKL-CH21) gene 8 exons and their homologous genes on chromosome 1 person muscular phosphofructokinase (the human muscle-type phosphofructokinase located on chromosome 1PFKM-CH1) exon 9, SYBR Green I dye PCR products, agar sugar gel electrophoresis, gel imaging system analysis software Quantitative One optical density, according to the amplified product PFKL-CH21/PFKM-CH1 optical density ratio for quantitative diagnosis. 2. Detection DS: ① The chromosomal G-banding karyotype analysis confirmed 26 cases of children with DS case group, 20 cases of normal control group; peripheral blood extracted DNA, stored in -20 ℃ spare. ② with SYBR Green I HGQ-PCR method for detection of DS and normal DNA. 3 Comparison of various detection methods: DNA template in the same patient, the same PCR reaction, the PCR products of different cycles, respectively, with 2% agarose gel electrophoresis, SYBR Green I dye, bromide has ingot (ethidium bromide, EB) staining, polyacrylamide gel electrophoresis - silver staining (referred to silver staining) to detect a variety of methods to compare the advantages and disadvantages. Results: 1.SYBR Green I HGQ-PCR method for detection of amplified products PFKL-CH21/PFKM-CH1 optical density ratio: The patient group was 1.61 ± 0.18 (x lt; WP = 4 gt; ± s); while the normal group 1.01 ± 0.06 (x ± s), by statistical analysis, two samples were compared, P lt; 0.01, the results consistent with karyotype analysis, about 4h can be completed. 2 test results of three methods: SYBR Green I dye to detect than EB staining sensitive, accurate and safe; same with silver staining sensitivity than silver staining is simple, fast, time-saving. Conclusion: 1.SYBR Green I homologous gene quantitative PCR-saving, simple, fast, and prenatal diagnosis for clinical provides a new diagnostic method. 2 stained with SYBR Green I fluorescence quantitative PCR method of homologous genes, can accurately diagnose DS, and karyotype analysis results. 3.SYBR Green I dye and silver staining equally sensitive enough to detect low concentrations, the low number of cycles DNA PCR product, and time-saving, fast. 4 a pair of primers amplifying two homologous genes simultaneously, eliminating a number of factors influence the PCR amplification, improves the accuracy quantitative PCR.
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