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首页> 外文期刊>Pharmacogenetics and genomics >A multiplex MALDI-TOF MS approach facilitates genotyping of DNA from formalin-fixed paraffin-embedded tumour specimens.
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A multiplex MALDI-TOF MS approach facilitates genotyping of DNA from formalin-fixed paraffin-embedded tumour specimens.

机译：多重MALDI-TOF MS方法可促进福尔马林固定石蜡包埋的肿瘤标本中DNA的基因分型。

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摘要

OBJECTIVE: The impact of single-nucleotide polymorphisms (SNPs) on tumour susceptibility and pathogenesis has gained enormous attention. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS)-based genotyping facilitates the analysis of short DNA amplicons and is, therefore, a promising tool for the investigation of formalin-fixed paraffin-embedded (FFPE) tissue samples, particularly in targeted genotyping analysis. METHODS: To examine the applicability of genotyping FFPE DNA with MALDI-TOF MS in multiplex reactions, we investigated five DNA samples extracted from FFPE tumour specimens from follicular lymphoma patients using different extraction methods (phenol-chloroform, commercial kit). Thirty-one SNPs from 25 genes, integrated in different-sized multiplex assays (7-plex, 10-plex, 14-plex, 24-plex), were analyzed. To investigate the reliability of genotyping tumour-derived DNA extracted from FFPE tissue, we examined 64 FFPE tumour specimens in comparison with matched germline DNA samples. RESULTS: Call rates of 99.6 (274/275) and 93.5% (257/275) were observed for the DNA extracted with the phenol-chloroform approach or the commercial extraction kit, respectively. Increasing the number of SNPs per assay resulted in reduced genotyping call rates and genotyping quality, especially in the DNA samples isolated with the commercial extraction kit. When comparing the genotypes of DNA derived from germline and tumour (FFPE) specimens, a perfect concordance rate of 100% was detected. CONCLUSION: Our data delineate that MALDI-TOF-based genotyping of FFPE DNA is reliable and reproducible even in multiplex reactions, enabling the retrospective investigation of FFPE study cohorts in future experiments.
机译：目的：单核苷酸多态性（SNPs）对肿瘤易感性和发病机制的影响已引起广泛关注。基于基质辅助的激光解吸/电离飞行时间质谱（MALDI-TOF MS）的基因分型有助于短DNA扩增子的分析，因此是研究福尔马林固定石蜡包埋（FFPE）的有前途的工具）组织样本，尤其是在靶向基因分型分析中。方法：为了研究利用MALDI-TOF MS对FFPE DNA进行基因分型在多重反应中的适用性，我们调查了使用不同提取方法（酚-氯仿，商业试剂盒）从滤泡性淋巴瘤患者的FFPE肿瘤标本中提取的5个DNA样品。分析了来自25个基因的31个SNP，这些基因已整合到不同大小的多重测定中（7重，10重，14重，24重）。为了调查从FFPE组织提取的肿瘤衍生DNA的基因型的可靠性，我们与匹配的种系DNA样品进行了比较，检查了64个FFPE肿瘤标本。结果：用苯酚-氯仿法或商业提取试剂盒提取的DNA的检出率分别为99.6（274/275）和93.5％（257/275）。每次测定中增加SNP的数量会导致基因分型检出率和基因分型质量降低，尤其是在使用商业提取试剂盒分离的DNA样品中。当比较来自种系和肿瘤（FFPE）标本的DNA的基因型时，检测到100％的完美一致性。结论：我们的数据表明，即使在多重反应中，基于MALDI-TOF的FFPE DNA的基因分型也是可靠且可重现的，从而可以对FFPE研究队列进行回顾性研究。

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