Global Sequencing approach for characterizing the molecular background of hereditary iron disorders
Global Sequencing approach for characterizing the molecular background of hereditary iron disorders
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DOI：
10.1373/clinchem.2007.090605
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发表时间：
2007-12-01
影响因子：
9.3
通讯作者：
Aguilar-Martinez, Patricia
中科院分区：
文献类型：
--
作者：
Cunat, Severine;Giansily-Blaizot, Muriel;Aguilar-Martinez, Patricia
背景：遗传性血色素沉着症（HH）或遗传性高铁蛋白血症（HF）的新遗传形式在过去几年中已被发现，并且在单个患者中各种基因的异常可能相互作用。本研究旨在开发一种快速的自动化方法来测序主要基因。方法：采用SCAIP （single-condition amplification with internal引物单条件扩增）方法，采用标准96孔微孔板，对HFE（血色素沉着症）、HAMP （hepcidin抗菌肽）、HFE2/HJV（2型血色素沉着症（幼型））、SLC40A1（铁转运蛋白）和TFR2（转铁蛋白受体2）基因以及FTL（铁蛋白，轻多肽）基因的5'未翻译区进行测序。为了进一步简化方法，我们调整了PCR条件以避免使用内部引物，并将这种单条件扩增方法应用于38例选择的无关联患者。我们根据临床情况量身定制了基因调查，患者分为两组。组1包括高铁素血症和高转铁蛋白饱和度（TS）的患者（典型的成人和青少年HH形式，分别为1A和1B组），组2包括高铁素血症和低、典型或轻微增加的TS，有或没有铁超载的患者（分别为2A和2B组）。结果：通过这种策略，我们确定了单基因和多基因异常，包括6个先前未描述的HFE (c.794dupA), HFE2 （c. 89-4duff）和SLC40A1 （c.262A>G, c.533G>A, c.1468G>A和c. 59_45del）异常。结论：该方法是研究遗传性铁超载或HF的简单方法，可快速评估患者。(C) 2007年美国临床化学学会。
Background: New genetic forms of hereditary hemochromatosis (HH) or hereditary hyperferritinemia (HF) have been identified over the last few years, and abnormalities of various genes may interact in a single patient. This study aimed to develop a rapid automated method for sequencing the main genes involved.Methods: We used a standard 96-well microplate with a single PCR condition in an adaptation of the SCAIP (single-condition amplification with internal primer) method to sequence the HFE (hemochromatosis), HAMP (hepcidin antimicrobial peptide), HFE2/HJV [hemochromatosis type 2 (juvenile)], SLC40A1 (ferroportin), and TFR2 (transferrin receptor 2) genes, and the 5' untranslated region of the FTL (ferritin, light polypeptide) gene. To further simplify the method, we adjusted PCR conditions to avoid the use of an internal primer and applied this single-condition amplification method to 38 selected, unrelated patients. We tailored the genetic investigation according to the clinical picture, with the patients falling into 2 groups. Group 1 consisted of patients with hyperferritinemia and high transferrin saturation (TS) (classic adult and juvenile HH forms, groups 1A and 1B, respectively), and group 2 consisted of patients with hyperferritinemia and low, typical, or slightly increased TS, with or without iron overload (groups 2A and 2B, respectively).Results: With this strategy we identified single-gene and multigene abnormalities, including 6 previously undescribed abnormalities in HFE (c.794dupA), HFE2 (c.-89-4duff), and SLC40A1 (c.262A>G, c.533G>A, c.1468G>A, and c.-59_-45del).Conclusion: This method is a simple approach for investigating hereditary iron overload or HF and allows rapid evaluation of patients. (C) 2007 American Association for Clinical Chemistry.