Detection of hemi/homozygotes through heteroduplex formation in high-resolution melting analysis
ABSTRACT Heteroduplex formation, required for the complete detection of hemi/homozygotes using high-resolution melting analysis, can be induced either by pre-PCR mixing of genomic DNAs or by post-PCR mixing of PCR products from unknown and reference samples. This study investigates the effects of both methods using two single nucleotide polymorphisms in X-linked DMD gene. The results show that both methods resulted in the same effect when mixing samples with the same gene copy number. Mixing samples with different gene copy numbers has not been previously explored and we show that post-PCR mixing is insensitive to gene copy number differences as compared to pre-PCR mixing.
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ABSTRACT: Nurr1 defects could in part underlie Parkinson's disease pathogenesis, and Nurr1 gene polymorphism has been found in Caucasian patients with Parkinson's disease. In this study, heteroduplex technology was applied to compare the DNA sequences of eight exons of Nurr1 among 200 sporadic Parkinson's disease patients and 200 healthy controls in the Han population in the Hubei province, China. One allele amplified from exon 3 of Nurr1 was polymorphic in five Parkinson's disease patients (2.5%, 5/200), and two individuals had a polymorphic allele amplified from exon 2 (1%, 2/200). The anomalous electrophoresis fragment in exon 3 of Nurr1 gene contained a 709C/A missense mutation, and a polymorphic single nucleotide polymorphism at 388G/A was identified in exon 2. Compared with the control group, the Nurr1 gene expression level in the Parkinson's disease group was decreased, and the Nurr1 gene expression levels in Parkinson's disease patients carrying the polymorphisms at exons 2 and 3 were significantly decreased. Our data indicate that the single nucleotide polymorphism 388G/A in exon 2 and the 709C/A missense mutation in exon 3 of the Nurr1 gene in the Chinese population might affect the pathogenesis of Parkinson's disease.Neural Regeneration Research 08/2012; 7(23):1791-6. DOI:10.3969/j.issn.1673-5374.2012.23.005 · 0.23 Impact Factor