Development of a 950-gene DNA array for examining gene expression patterns in mouse testis

BACKGROUND: Over the past five years, interest in and use of DNA array technology has increased dramatically, and there has been a surge in demand for different types of arrays. Although manufacturers offer a number of pre-made arrays, these are generally of utilitarian design and often cannot accommodate the specific requirements of focused research, such as a particular set of genes from a particular tissue. We found that suppliers did not provide an array to suit our particular interest in testicular toxicology, and therefore elected to design and produce our own. RESULTS: We describe the procedures used by members of the US Environmental Protection Agency MicroArray Consortium (EPAMAC) to produce a mouse testis expression array on both filter and glass-slide formats. The approaches used in the selection and assembly of a pertinent, nonredundant list of testis-expressed genes are detailed. Hybridization of the filter arrays with normal and bromochloroacetic acid-treated mouse testicular RNAs demonstrated that all the selected genes on the array were expressed in mouse testes. CONCLUSION: We have assembled two lists of mouse (950) and human (960) genes expressed in the mouse and/or human adult testis, essentially all of which are available as sequence-verified clones from public sources. Of these, 764 are homologous and will therefore enable close comparison of gene expression between murine models and human clinical testicular samples.

背景：过去五年间，DNA微阵列（DNA array）技术的关注度与应用规模均大幅提升，各类微阵列的市场需求也呈现爆发式增长。尽管厂商推出了多款预制微阵列产品，但这类产品普遍采用通用化设计，往往无法满足针对性研究的特定需求，例如针对特定组织的特定基因集。我们发现，现有供应商未提供适配我们睾丸毒理学研究方向的微阵列，因此决定自行设计并制备此类微阵列。
结果：本文详述了美国环境保护署微阵列联盟（US Environmental Protection Agency MicroArray Consortium, EPAMAC）成员所采用的流程，分别制备了滤膜式与玻片式两种格式的小鼠睾丸表达谱微阵列。同时详细说明了筛选并组装非冗余且贴合研究主题的睾丸表达基因列表的方法。将滤膜微阵列与正常小鼠睾丸RNA、溴氯乙酸（bromochloroacetic acid）处理后的小鼠睾丸RNA进行杂交实验，结果显示阵列上的所有选定基因均在小鼠睾丸中表达。
结论：我们已分别构建了小鼠（950个）与人类（960个）在成年睾丸中表达的基因列表，其中绝大多数均可从公共数据源获取经过序列验证的克隆。在这些基因中，有764个为同源基因，因此可用于精准比较小鼠模型与人类临床睾丸样本之间的基因表达差异。