Kringel Dario, Kaunisto Mari A, Lippmann Catharina, Kalso Eija, Lötsch Jörn
Institute of Clinical Pharmacology, Goethe-University, Frankfurt, Germany.
Institute for Molecular Medicine Finland, HiLIFE, University of Helsinki, Helsinki, Finland.
Front Pharmacol. 2018 Sep 19;9:1008. doi: 10.3389/fphar.2018.01008. eCollection 2018.
Many gene variants modulate the individual perception of pain and possibly also its persistence. The limited selection of single functional variants is increasingly being replaced by analyses of the full coding and regulatory sequences of pain-relevant genes accessible by means of next generation sequencing (NGS). An NGS panel was created for a set of 77 human genes selected following different lines of evidence supporting their role in persisting pain. To address the role of these candidate genes, we established a sequencing assay based on a custom AmpliSeq panel to assess the exomic sequences in 72 subjects of Caucasian ethnicity. To identify the systems biology of the genes, the biological functions associated with these genes were assessed by means of a computational over-representation analysis. Sequencing generated a median of 2.85 ⋅ 10 reads per run with a mean depth close to 200 reads, mean read length of 205 called bases and an average chip loading of 71%. A total of 3,185 genetic variants were called. A computational functional genomics analysis indicated that the proposed NGS gene panel covers biological processes identified previously as characterizing the functional genomics of persisting pain. Results of the NGS assay suggested that the produced nucleotide sequences are comparable to those earned with the classical Sanger sequencing technique. The assay is applicable for small to large-scale experimental setups to target the accessing of information about any nucleotide within the addressed genes in a study cohort.
许多基因变异会调节个体对疼痛的感知，也可能影响疼痛的持续时间。以往对单个功能变异的有限选择正逐渐被对疼痛相关基因的完整编码和调控序列的分析所取代，这些分析可通过新一代测序（NGS）技术实现。我们根据支持其在持续性疼痛中作用的不同证据线索，挑选了77个人类基因，创建了一个NGS检测板。为了研究这些候选基因的作用，我们基于定制的扩增子测序检测板建立了一种测序分析方法，以评估72名白种人受试者的外显子序列。为了确定这些基因的系统生物学特性，我们通过计算性过度表达分析评估了与这些基因相关的生物学功能。每次测序运行产生的读数中位数为2.85×10，平均深度接近200次读数，平均读长为205个已识别碱基，芯片平均加载率为71%。总共识别出3185个基因变异。计算功能基因组学分析表明，所提出的NGS基因检测板涵盖了先前确定的作为持续性疼痛功能基因组学特征的生物学过程。NGS分析结果表明，所产生的核苷酸序列与经典桑格测序技术获得的序列相当。该检测方法适用于从小规模到大规模的实验设置，以获取研究队列中所研究基因内任何核苷酸的信息。
