Herbert John M J, Stekel Dov J, Mura Manuela, Sychev Michail, Bicknell Roy
Cancer Research UK Angiogenesis Group, Institute for Biomedical Research, Schools of Immunity and Infection and Cancer studies, College of Medicine and Dentistry, University of Birmingham, Birmingham, UK.
Methods Mol Biol. 2011;729:99-119. doi: 10.1007/978-1-61779-065-2_7.
The aim of this method is to guide a bench scientist to maximise cDNA library analyses to predict biologically relevant genes to pursue in the laboratory. Many groups have successfully utilised cDNA libraries to discover novel and/or differentially expressed genes in pathologies of interest. This is despite the high cost of cDNA library production using the Sanger method of sequencing, which produces modest numbers of expressed sequences compared to the total transcriptome. Both public and propriety cDNA libraries can be utilised in this way, and combining biologically relevant data can reveal biologically interesting genes. Pivotal to the quality of target identification are the selection of biologically relevant libraries, the accuracy of Expressed Sequence Tag to gene assignment, and the statistics used. The key steps, methods, and tools used to this end will be described using vascular targeting as an example. With the advent of next-generation sequencing, these or similar methods can be applied to find novel genes with this new source of data.
该方法的目的是指导实验台科学家最大限度地进行cDNA文库分析,以预测在实验室中值得研究的具有生物学相关性的基因。许多研究小组已成功利用cDNA文库在感兴趣的病理学中发现新的和/或差异表达的基因。尽管使用桑格测序法生产cDNA文库成本高昂,且与整个转录组相比,其产生的表达序列数量有限。公共和专用的cDNA文库均可按此方式使用,整合生物学相关数据能够揭示具有生物学意义的基因。目标识别质量的关键在于生物学相关文库的选择、表达序列标签到基因分配的准确性以及所使用的统计方法。为此,将以血管靶向为例描述关键步骤、方法和工具。随着下一代测序技术的出现,这些或类似方法可应用于利用这一新数据源寻找新基因。