Data from: Double-digest RAD Sequencing using Ion Proton semiconductor platform (ddRADseq-ion) with non-model organisms

Research in evolutionary biology involving nonmodel organisms is rapidly shifting from using traditional molecular markers such as mtDNA and microsatellites to higher throughput SNP genotyping methodologies to address questions in population genetics, phylogenetics and genetic mapping. Restriction site associated DNA sequencing (RAD sequencing or RADseq) has become an established method for SNP genotyping on Illumina sequencing platforms. Here, we developed a protocol and adapters for double-digest RAD sequencing for Ion Torrent (Life Technologies; Ion Proton, Ion PGM) semiconductor sequencing. We sequenced thirteen genomic libraries of three different nonmodel vertebrate species on Ion Proton with PI chips: Arctic charr Salvelinus alpinus, European whitefish Coregonus lavaretus and common lizard Zootoca vivipara. This resulted in ~962 million single-end reads overall and a mean of ~74 million reads per library. We filtered the genomic data using Stacks, a bioinformatic tool to process RAD sequencing data. On average, we obtained ~11 000 polymorphic loci per library of 6–30 individuals. We validate our new method by technical and biological replication, by reconstructing phylogenetic relationships, and using a hybrid genetic cross to track genomic variants. Finally, we discuss the differences between using the different sequencing platforms in the context of RAD sequencing, assessing possible advantages and disadvantages. We show that our protocol can be used for Ion semiconductor sequencing platforms for the rapid and cost-effective generation of variable and reproducible genetic markers.

针对非模式生物的进化生物学研究，正快速从线粒体DNA（mitochondrial DNA, mtDNA）、微卫星等传统分子标记，转向高通量单核苷酸多态性（Single Nucleotide Polymorphism, SNP）基因分型技术，以解答群体遗传学、系统发育学与遗传作图领域的科学问题。限制性酶切位点相关DNA测序（Restriction site associated DNA sequencing, 简称RAD测序或RADseq）已成为Illumina测序平台上成熟的SNP基因分型方法。本研究针对Ion Torrent（Life Technologies，即生命科技公司；旗下包含Ion Proton、Ion PGM）半导体测序平台，开发了双酶切RAD测序的实验方案与接头序列。我们使用PI芯片在Ion Proton平台上，对3种不同非模式脊椎动物的13个基因组文库进行了测序：北极红点鲑（Salvelinus alpinus）、欧洲白鲑（Coregonus lavaretus）与胎生蜥蜴（Zootoca vivipara）。最终共获得约9.62亿条单端测序读段（single-end reads），单个文库的平均读段数约为7400万条。我们借助生物信息学工具Stacks对基因组数据进行过滤处理。平均而言，每个包含6~30个个体的文库可获得约11000个多态性位点。我们通过技术重复与生物学重复、重构系统发育关系，以及利用杂交遗传群体追踪基因组变异的方式，验证了新方法的可靠性。最后，我们探讨了RAD测序中不同测序平台的应用差异，评估了各自的潜在优势与不足。结果表明，本研究开发的方案可应用于Ion半导体测序平台，实现快速且经济高效地生成兼具多态性与可重复性的遗传标记。