ARUP Institute for Clinical and Experimental Pathology, University of Utah Health Sciences Center, Salt Lake City, UT 84108, USA.
Hum Mutat. 2010 May;31(5):588-93. doi: 10.1002/humu.21230.
Germline mutation detection in PMS2, one of four mismatch repair genes associated with Lynch syndrome, is greatly complicated by the presence of numerous pseudogenes. We used a modification of a long-range PCR method to evaluate PMS2 in 145 clinical samples. This modification avoids potential interference from the pseudogene PMS2CL by utilizing a long-range product spanning exons 11-15, with the forward primer anchored in exon 10, an exon not shared by PMS2CL. Large deletions were identified by MLPA. Pathogenic PMS2 mutations were identified in 22 of 59 patients whose tumors showed isolated loss of PMS2 by immunohistochemistry (IHC), the IHC profile most commonly associated with a germline PMS2 mutation. Three additional patients with pathogenic mutations were identified from 53 samples without IHC data. Thirty-seven percent of the identified mutations were large deletions encompassing one or more exons. In 27 patients whose tumors showed absence of either another protein or combination of proteins, no pathogenic mutations were identified. We conclude that modified long-range PCR can be used to preferentially amplify the PMS2 gene and avoid pseudogene interference, thus providing a clinically useful germline analysis of PMS2. Our data also support the use of IHC screening to direct germline testing of PMS2.
胚系突变检测在 PMS2 中，四个错配修复基因与林奇综合征相关的一个，由于存在许多假基因而变得非常复杂。我们使用一种长距离 PCR 方法的改进来评估 145 个临床样本中的 PMS2。这种改进通过利用跨越外显子 11-15 的长距离产物来避免假基因 PMS2CL 的潜在干扰，正向引物锚定在外显子 10 上，该外显子与 PMS2CL 不共享。通过 MLPA 鉴定大片段缺失。通过免疫组织化学（IHC）显示 PMS2 孤立缺失的 59 名患者中的 22 名患者中鉴定出致病性 PMS2 突变，IHC 图谱最常与胚系 PMS2 突变相关。从 53 个没有 IHC 数据的样本中另外鉴定出 3 名具有致病性突变的患者。鉴定出的突变中有 37%是包含一个或多个外显子的大片段缺失。在 27 名肿瘤显示另一种蛋白或蛋白组合缺失的患者中，未鉴定出致病性突变。我们得出结论，改良的长距离 PCR 可用于优先扩增 PMS2 基因并避免假基因干扰，从而提供有用的 PMS2 胚系分析。我们的数据还支持使用 IHC 筛选来指导 PMS2 的种系测试。
