Ullate-Agote Asier, Chan Yingguang Frank, Tzika Athanasia C
Laboratory of Artificial and Natural Evolution (LANE), Department of Genetics and Evolution, University of Geneva, Sciences III, 30, Quai Ernest-Ansermet, 1211, Geneva, Switzerland.
SIB Swiss Institute of Bioinformatics, Geneva, Switzerland.
Methods Mol Biol. 2017;1650:47-67. doi: 10.1007/978-1-4939-7216-6_3.
Multiple technologies and software are now available facilitating the de novo sequencing and assembly of any vertebrate genome. Yet the quality of most available sequenced genomes is substantially poorer than that of the golden standard in the field: the human genome. Here, we present a step-by-step protocol for the successful sequencing and assembly of a high-quality snake genome that can be applied to any other reptilian or avian species. We combine the great sequencing depth and accuracy of short reads with the use of different insert size libraries for extended scaffolding followed by optical mapping. We show that this procedure improved the corn snake scaffold N50 from 3.7 kbp to 1.4 Mbp, currently making it one of the snake genomes with the longest scaffolds. Short guidelines are also given on the extraction of long DNA molecules from reptilian blood and the necessary modifications in DNA extraction protocols. This chapter is accompanied by a website ( www.reptilomics.org/stepbystep.html ), where we provide links to the suggested software, examples of input and output files, and running parameters.
现在有多种技术和软件可用于促进对任何脊椎动物基因组的从头测序和组装。然而，大多数现有测序基因组的质量比该领域的黄金标准——人类基因组——要差得多。在此，我们展示了一个用于成功测序和组装高质量蛇基因组的分步方案，该方案可应用于任何其他爬行动物或鸟类物种。我们将短读长测序的高深度和准确性与使用不同插入片段大小文库进行扩展支架构建以及光学图谱相结合。我们表明，此程序将玉米蛇的支架N50从3.7千碱基对提高到了1.4兆碱基对，使其目前成为具有最长支架的蛇基因组之一。还给出了从爬行动物血液中提取长DNA分子的简短指南以及DNA提取方案中的必要修改。本章还配有一个网站（www.reptilomics.org/stepbystep.html），我们在该网站上提供了建议软件的链接、输入和输出文件示例以及运行参数。
