Development of a new genotyping assay for detection of the BDNFVal66Met polymorphism using melting-curve analysis
Development of a new genotyping assay for detection of the BDNFVal66Met polymorphism using melting-curve analysis
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DOI：
10.2217/pgs.09.44
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发表时间：
2009-06-01
期刊：
影响因子：
2.1
通讯作者：
Llerena, Adrian
中科院分区：
文献类型：
--
作者：
Antonia Sanchez-Romero, Maria;Dorado, Pedro;Llerena, Adrian
脑源性神经营养因子（BDNF）在中枢神经系统神经元的生长、分化和存活中起关键作用。最近的研究表明，BDNF可能与心境障碍和精神分裂症的病因有关，也与一些药物的治疗作用有关，如抗抑郁药和抗精神病药。本研究旨在建立一种简单、快速、准确的新方法，应用DNA特异性染料SYBR（R）绿色I和熔解曲线分析法检测精神分裂症患者BDNF基因Val66 Met多态性。应用SYBR绿色I标记的PCR产物荧光熔解曲线分析法，对30例精神分裂症患者进行BDNF Val66 Met多态性（rs6265）基因分型。使用特异性BDNF TaqMan（R）等位基因辨别测定来确认所有30个样品的基因型结果。这种新方法允许使用SYBR绿色1在同一反应管中分析两个等位基因，而不需要额外的步骤。GC钳的添加使得该方法普遍适用，因为可以根据需要调节一个等位基因的解链温度以给出解链曲线的独特分离。因此，该方法简便、快速、准确，可用于BDNF Val66Met多态性的检测。它也可能是有用的药物遗传学研究中的其他SNPs的分析。
Brain-derived neurotrophic factor (BDNF) plays a critical role in the growth, differentiation and survival of neurons in the CNS. Recent research has suggested that BDNF may be implicated in the etiology of mood disorders and schizophrenia, as well as in the therapeutic action of some drugs, such as antidepressants and antipsychotics. This study aimed to develop a simple, fast and accurate new method for detecting the Val66Met polymorphism of the BDNF gene in schizophrenia patients using melting-curve analysis and a DNA-specific dye, SYBR (R) Green I. A group of 30 schizophrenia patients were analyzed to detect the BDNF Val66Met polymorphism (rs6265) using the new genotyping method based on the analysis of fluorescence melting curves of PCR products that were labeled with SYBR Green I. The genotype results were confirmed for all 30 samples using the specific BDNF TaqMan (R) allele discrimination assay. This new method allows the analysis of both alleles in the same reaction tube using SYBR Green 1, with no need for additional steps. The addition of a GC clamp makes this method universally applicable, since the melting temperature of one allele can be adjusted as necessary to give the distinctive separation of melting curves. Therefore, this new method is simple, fast and accurate for determining the presence of the BDNF Val66Met polymorphism. It may also be useful for the analysis of other SNPs in pharmacogenetic studies.